Connect public, paid and private patent data with Google Patents Public Datasets

Synthetic antigen for the detection of aids-related disease

Download PDF

Info

Publication number
US20030211117A1
US20030211117A1 US10305271 US30527102A US2003211117A1 US 20030211117 A1 US20030211117 A1 US 20030211117A1 US 10305271 US10305271 US 10305271 US 30527102 A US30527102 A US 30527102A US 2003211117 A1 US2003211117 A1 US 2003211117A1
Authority
US
Grant status
Application
Patent type
Prior art keywords
leu
glu
gln
peptide
ala
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Abandoned
Application number
US10305271
Inventor
Wesley Cosand
Original Assignee
Cosand Wesley Loren
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date

Links

Images

Classifications

    • CCHEMISTRY; METALLURGY
    • C07ORGANIC CHEMISTRY
    • C07KPEPTIDES
    • C07K14/00Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/005Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from viruses
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09Recombinant DNA-technology
    • C12N15/63Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/66General methods for inserting a gene into a vector to form a recombinant vector using cleavage and ligation; Use of non-functional linkers or adaptors, e.g. linkers containing the sequence for a restriction endonuclease
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL, OR TOILET PURPOSES
    • A61K38/00Medicinal preparations containing peptides
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL, OR TOILET PURPOSES
    • A61K39/00Medicinal preparations containing antigens or antibodies
    • CCHEMISTRY; METALLURGY
    • C07ORGANIC CHEMISTRY
    • C07KPEPTIDES
    • C07K2319/00Fusion polypeptide
    • CCHEMISTRY; METALLURGY
    • C07ORGANIC CHEMISTRY
    • C07KPEPTIDES
    • C07K2319/00Fusion polypeptide
    • C07K2319/01Fusion polypeptide containing a localisation/targetting motif
    • C07K2319/02Fusion polypeptide containing a localisation/targetting motif containing a signal sequence
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2740/00Reverse Transcribing RNA Viruses
    • C12N2740/00011Reverse Transcribing RNA Viruses
    • C12N2740/10011Retroviridae
    • C12N2740/16011Human Immunodeficiency Virus, HIV
    • C12N2740/16111Human Immunodeficiency Virus, HIV concerning HIV env
    • C12N2740/16122New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2740/00Reverse Transcribing RNA Viruses
    • C12N2740/00011Reverse Transcribing RNA Viruses
    • C12N2740/10011Retroviridae
    • C12N2740/16011Human Immunodeficiency Virus, HIV
    • C12N2740/16211Human Immunodeficiency Virus, HIV concerning HIV gagpol
    • C12N2740/16222New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
    • YGENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10STECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S435/00Chemistry: molecular biology and microbiology
    • Y10S435/961Chemistry: molecular biology and microbiology including a step of forming, releasing, or exposing the antigen or forming the hapten-immunogenic carrier complex or the antigen per se
    • YGENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10STECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S435/00Chemistry: molecular biology and microbiology
    • Y10S435/974Aids related test
    • YGENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10STECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S930/00Peptide or protein sequence
    • Y10S930/01Peptide or protein sequence
    • Y10S930/22Viral peptide or viral protein
    • Y10S930/221Retrovirus related, or human immunodeficiency virus related, or simian immunodeficiency virus related

Abstract

Novel peptides are provided having substantially the same sequence as immunologically significant fragments of AIDS-related viruses. The polypeptides can be used as reagents in the determination of exposure of a human host to the virus. Of particular interest is the use of polypeptides in screening blood products.

Description

    CROSS-REFERENCE TO RELATED APPLICATIONS
  • [0001]
    This application is a Continuation-in-part of U.S. application Ser. No. 767,303, filed Aug. 19, 1985 which is a Continuation-in-part of U.S. application Ser. No. 728,052, filed Apr. 29, 1985, which disclosures are incorporated herein by reference.
  • BACKGROUND OF THE INVENTION Field of the Invention
  • [0002]
    With the discovery that the diseases called lymphadenopathy syndrome and acquired immune deficiency disease (AIDS) are caused by an infectious retrovirus designated lymphadenopathy virus (LAV), human T-cell lymphotropic virus-III (HTLV-III), AIDS-related virus (ARV), or immune deficiency-associated virus (IDAV), there has become an immediate need to be able to detect potential vectors of the disease, such as blood from diseased individuals, which may be employed for transfusions or from which specific blood factors may be isolated.
  • [0003]
    To detect potential vectors of the disease, it is necessary to have viral proteins and/or antibodies to such proteins. Because of the hazards associated with growing the LAV/HTLV-III retrovirus, there is significant interest in establishing means for obtaining the viral proteins or their immunologic equivalents, which means do not necessitate handling large volumes of live, potentially infectious virus. In choosing alternatives, one must be concerned with the fact that the viruses have been reported to be highly polymorphic, frequently changing as the retrovirus is passaged.
  • BRIEF DESCRIPTION OF THE RELEVANT LITERATURE
  • [0004]
    The various antigens of the retrovirus are described by Saxinger et al., Science (1985) 227:1036-1038. See also Gallo et al., ibid. (1984) 224:500; Sarangadharn et al., ibid. 224:506; Barre-Sinoussi et al., ibid. (1983) 220:868; Montagnier et al., in Human T-Cell Leukemia/Lymphoma Virus, Gallo, Essex, Gross, eds. (Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y.), 1984, p. 363. These may include, but are not limited to, p13, p18, p25, p36, gp43, p55, gp65, gp110, etc., where the numbers may differ depending upon the reporter.
  • [0005]
    Hopp and Woods, Proc. Natl. Acad. Sci. USA (1981) 78:3824, describe criteria for selecting peptides as potential epitopes of polypeptides based on their relative hydrophilicity. In one study employing these criteria, a 12-amino acid peptide was synthesized that bound 9% of antibodies elicited by the native protein (Hopp, Molec. Immunol. (1981) 18:869). In general, Hopp/Woods criteria have been shown not to have a high predictive value. Furthermore, epitopes have been demonstrated which are not hydrophilic (Kazim et al., Biochem. J. (1982) 203:201). Other studies of polypeptide antigenicity include Green et al., Cell (1982) 28:477, where peptides were employed which elicited antibodies, which antibodies were capable of binding to the native protein, while conversely antibodies which were elicited by the native protein failed to bind to the peptides; and Trainer et al., Nature (1984) 312:127, whose results with myohaemerythrin paralleled those of Green et al.
  • [0006]
    The complete nucleotide sequence of LAV is reported by Wain-Hobson et al., Cell (1985) 40:9. The complete sequence for HTLV-III is reported by Muesing et al., Nature (1985) 313:450, while the complete sequence for ARV is reported by Sanchez-Pescador et al., Science (1985) 227:484. All three viruses exhibit substantial nucleotide homology and are similar with respect to morphology, cytopathology, requirements for optimum reverse transcriptase activity, and at least some antigenic properties (Levy et al., Science (1984) 225:840; Shupbach et al., Science (1984) 224:503), and hence should be considered isolates of the same virus. See also, Chang et al., Science (1985) 228:93.
  • SUMMARY OF THE INVENTION
  • [0007]
    Peptide sequences capable of immunologically mimicking proteins encoded in the gag and/or env regions of the LAV/HTLV-III retrovirus are provided as reagents for use in the screening of blood and blood products for prior exposure to the retrovirus. The peptides are of at least 5 amino acids and can be used in various specific binding assays for the detection of antibodies to LAV/HTLV-III virus, for the detection of LAV/HTLV-III antigens, or as immunogens.
  • DESCRIPTION OF THE SPECIFIC EMBODIMENTS
  • [0008]
    For the purpose of this disclosure, a virus is considered to be the same as or equivalent to LAV/HTLV-III if it substantially fulfills the following criteria:
  • [0009]
    (a) The virus is tropic for T-lymphocytes, especially T-helper cells (CD4, according to the international nomenclature defined in Bernard et al., eds. Leucocyte Typing, New York: Springer Verlag, 1984);
  • [0010]
    (b) The virus is cytopathic for infected CD4+ cells (rather than transforming, as are HTLV-I and -II);
  • [0011]
    (c) The virus encodes an RNA-dependent DNA polymerase (reverse transcriptase) which is Mg2+-dependent (optimum concentration 5 nM), has a pH optimum of 7.8, is not inhibitable by actinomycin D, and can employ oligo(dT)12-18 as a primer for reverse transcription from its 3′ LTR;
  • [0012]
    (d) The virus bands in a sucrose gradient at a density of approximately 1.16;
  • [0013]
    (e) The virus can be labeled with [3H]-uridine;
  • [0014]
    (f) The virus is substantially cross-reactive immunologically with the proteins encoded by the gag and env regions of LAV/HTLV-III; and
  • [0015]
    (g) The virus shares substantial nucleotide homology (approximately 75-100%) and amino acid sequence homology (approximately 75-100%) with LAV or HTLV-III.
  • [0016]
    Novel peptides are provided which immunologically mimic proteins encoded by the LAV/FITLV-III retrovirus, particularly proteins encoded by the gag and/or env regions of the viral genome. To accommodate strain-to-strain variations among different isolates, adjustments for conservative substitutions and selection among the alternatives where non-conservative substitutions are involved, may be made. These peptides can be used individually or together for detection of the virus or of antibodies to the virus in a physiological sample. Depending upon the nature of the test protocol, the peptides may be labeled or unlabeled, bound to a solid surface, conjugated to a carrier or other compounds, or the like.
  • [0017]
    The peptides of interest will be derived from the peptides encoded by the gag region or the env region. These peptides will be primarily derived from p55 or fragments thereof, e.g., p25 and p18, or gp150 and fragments thereof, e.g., gp41. These peptides will be given Roman numerals, but will also be given numerical designations which are arbitrarily associated with the manner in which they were produced.
  • [0018]
    For the gag region, of particular interest are the coding regions extending from about base pair (bp) 450 to bp 731, particularly from about bp 450 to bp 545 (97) and bp 696 to bp 731 (71); from about bp 900 to bp 1421, particularly from about bp 921 to bp 1016, including bp 921 to bp 1010; bp 972 to bp 1016 (92); and bp 936 to bp 995 (17); or from about bp 1158 to about bp 1400, particularly bp 1164 to bp 1250 (90); bp 1278 to bp 1385 (88); and bp 1320 to bp 1385 (15), of the LAV/HTLV-III retrovirus. (Numbering according to Wain-Hobson et al., supra.)
  • [0019]
    For the env region, the regions of particular interest will be those polypeptides encoded within the bp 7210 to bp 7815 regions, particularly within bp 7231 to bp 7794, more particularly within about bp 7246 through bp 7317 (36), bp 7516 through bp 7593 (39), particularly bp 7543 through bp 7593 (79) and bp 7561 through 7593 (78), bp 7708 through bp 7779 (23), bp 7630 through bp 7689 (40), bp 7498 through bp 7554 (56).
  • [0020]
    The peptides of interest will include at least five, sometimes six, sometimes eight, sometimes 12, usually fewer than about 50, more usually fewer than about 35, and preferably fewer than about 25 amino acids included within a sequence coded for by the LAV/HTLV-III retrovirus. In each instance, desirably the oligopeptide will be as small as possible, while still maintaining substantially all of the sensitivity of the larger peptide. In some instances it may be desirable to join two or more oligopeptides which are non-overlapping in the same peptide structure or as individual peptides, which separately or together provide equivalent sensitivity to the parent.
  • [0021]
    The peptides may be modified by introducing conservative or non-conservative substitutions in the peptides, usually fewer than 20 number percent, more usually fewer than 10 number percent of the peptides being exchanged. In those situations where regions are found to be polymorphic, it may be desirable to vary one or more particular amino acids to more effectively mimic the differing epitopes of the different retroviral strains. In many instances to provide chemical and physical stability, methionine may be replaced by norleucine (Nor).
  • [0022]
    It should be understood that the polypeptides employed in the subject invention need not be identical to any particular LAV/HTLV-III polypeptide sequence, so long as the subject compounds are able to provide for immunological competition with proteins of at least one of the strains of the LAV/HTLV-III retrovirus. Therefore, the subject polypeptides may be subject to various changes, such as insertions, deletions, and substitutions, either conservative or non-conservative, where such changes might provide for certain advantages in their use. By conservative substitutions is intended combinations such as gly, ala; val, ile, leu; asp, glu; asn, gln; ser, thr; lys, arg; and phe, tyr. Usually, the sequence will not differ by more than twenty percent from the sequence of at least one strain of an LAV/HTLV-III retrovirus except where additional amino acids may be added at either terminus for the purpose of providing an “arm” by which the peptides of this invention may be conveniently immobilized. The arms will usually be at least 1 amino acid and may be 50 or more amino acids, more often 1 to 10 amino acids.
  • [0023]
    In addition, one or two amino acids may be added to the termini of an oligopeptide or peptide to provide for ease of linking of peptides one to another, for coupling to a support or larger peptide, for reasons to be discussed subsequently, for modifying the physical or chemical properties of the peptide or oligopeptide, or the like.
  • [0024]
    Amino acids such as tyrosine, cysteine, lysine, glutamic or aspartic acid, or the like, may be introduced at the C- or N-terminus of the peptide or oligopeptide to provide for a useful functionality for linking. Of particular interest is the presence of from 1 to 3 cysteines at the C- or N-terminus for linking to a support. The cysteine could be linked through a disulfide linkage to a dithio- or thio-functionalized support on a thioether linkage to an activated olefin support.
  • [0025]
    In addition, the peptide or oligopeptide sequences may differ from the natural sequence by the sequence being modified by terminal-NH2 acylation, e.g., acetylation, or thioglycolic acid amidation, terminal-carboxy amidation, e.g., ammonia, methylamine, etc. In some instances, these modifications may provide sites for linking to a support or other molecule.
  • [0026]
    The peptides and oligopeptides of interest will now be considered. The first peptides of interest will be derived from the gag region, particularly the protein referred to as p25 and p18.
  • [0027]
    The peptides for p25 are as follows:
  • [0028]
    The peptide I (15) encoded in the region bp 1320 to bp 1385 will have the following amino acid sequence, where oligopeptides included within the following sequence will include linear epitopes within such sequence:
    (I) (15)
    Y-Asp-Cys-Lys-Thr-Ile-Leu-Lys-Ala-Leu-
    Gly-Pro-Ala-Ala-Thr-Leu-Glu-Glu-Met-Met-
    Thr-Ala-Cys-X, where X is OH or NH2,
  • [0029]
    wherein the amino terminal Y, e.g., Tyr or Cys, if present, is added to facilitate coupling of the peptide to a protein carrier.
  • [0030]
    The next peptide II (17) will be encoded by the region extending from about bp 936 to bp 995 and will have the following sequence, where oligopeptides included within the following sequence will include linear epitopes within such sequence:
    (II) (17)
    Y-Leu-Lys-Glu-Thr-Ile-Asn-Glu-Glu-Ala-
    Ala-Glu-Trp-Asp-Arg-Val-His-Pro-Val-His-
    Ala-X, where X is OH or NH2,
  • [0031]
    wherein the amino terminal Y has been defined previously.
  • [0032]
    Of particular interest is the oligopeptide IIa:
    (IIa)
    Y-Ala-Ala-Glu-Trp-Asp-Arg-Z-X,
  • [0033]
    wherein X and Y have been defined previously and Z is a bond, an amino acid which provides a means of linking, e.g., cysteine, tyrosine, etc., or taken together with X provides a functional group which may be used for linking, e.g., an olefin as in allyl or maleimidyl, dithio, etc.
  • [0034]
    The next peptide of interest, III (92), will be encoded by the region extending from about bp 972 to bp 1016 and will have the following sequence, where oligopeptides included within the following sequence will include linear epitopes within such sequence:
    (III) (92)
    Y-Asp-Arg-Val-His-Pro-Val-His-Ala-Gly-Pro-
    Ile-Ala-Pro-Gly-Gln-X,
  • [0035]
    wherein X, Y and Z have been defined previously.
  • [0036]
    Preferably, this peptide will have no more than about 15 amino acids encoded by the LAV/HTLV III genome.
  • [0037]
    The next peptide, IV (90), will be encoded by the region extending from about bp 1164 to bp 1250 and will have the following sequence, where oligopeptides included within the following sequence will include linear epitopes within such sequence:
    (IV) (90)
    Y-Tyr-Ser-Pro-Thr-Ser-Ile-Leu-Asp-Ile-Arg-
    Gln-Gly-Pro-Lys-Glu-Pro-Phe-Arg-Asp-Tyr-Val-
    Asp-Arg-Phe-Tyr-Lys-Thr-Leu-Arg-Z-X,
  • [0038]
    wherein X, Y and Z have been defined previously.
  • [0039]
    Preferably, this peptide will have no more than about 29 amino acids encoded by the LAV/HTLV III genome.
  • [0040]
    The peptide, V (88), will be encoded by the region extending from about bp 1278 to bp 1385 and will have the following sequence, where oligopeptides included within the following sequence will include linear epitopes within such sequence:
    (V) (88)
    Y-Asn-Trp-Nor-Thr-Glu-Thr-Leu-Leu-Val-Gln-
    Asn-Ala-Asn-Pro-Asp-Cys-Lys-Thr-Ile-Leu-Lys-
    Ala-Leu-Gly-Pro-Ala-Ala-Thr-Leu-Glu-Glu-Nor-
    Nor-Thr-Ala-Cys-X,
  • [0041]
    wherein X and Y have been defined previously.
  • [0042]
    The next peptides of interest will be derived from the gag protein region referred to as p18.
  • [0043]
    The next peptide of interest, VI (97), will be encoded by the region extending from about bp 450 through bp 545 and will have the following sequence, where oligopeptides included within the following sequence will include linear epitopes within such sequence:
    (VI) (97)
    Y-Arg-Glu-Leu-Glu-Arg-Phe-Ala-Val-Asn-Pro-Gly-
    Leu-Leu-Glu-Thr-Ser-Glu-Gly-Cys-Arg-Gln-Ile-
    Leu-Gly-Gln-Leu-Gln-Pro-Ser-Leu-Gln-Thr-X,
  • [0044]
    wherein X and Y have been defined previously.
  • [0045]
    The next peptide of interest VII (71) will be encoded by the region extending from about bp 696 to bp 731. This peptide will include any oligopeptides coding for linear epitopes with the following amino acid sequence:
    (VII) (71)
    Y-Asp-Thr-Gly-His-Ser-Ser-Gln-Val-Ser-Gln-
    Asn-Tyr,
  • [0046]
    wherein Y has been defined previously.
  • [0047]
    The next polypeptides of interest will be those derived from the env region, from gp110 (110 kDal).
  • [0048]
    The next peptide of interest, VIII, will be encoded by the region extending from about bp 7246 through bp 7317 and while coming within the general limitations indicated previously, will preferably have no more than 24 amino acids encoded by the LAV/HTLV III genome.
  • [0049]
    The peptide of interest will generally have the following amino acid sequence, where oligopeptides included within the following sequence will include linear epitopes within such sequence:
    (VIII) (36)
    Val-Lys-Ile-Glu-Pro-Leu-Gly-Val-Ala-Pro-
    Thr-Lys-Ala-Lys-Arg-Arg-Val-Val-Gln-Arg-
    Glu-Lys-Arg-Ala-Z-X, where X is OH or NH2,
  • [0050]
    wherein the carboxy terminal Z, e.g., Cys, if present, is an amino acid added to facilitate coupling of the peptide to a protein carrier.
  • [0051]
    Of particular interest is where 6, conveniently up to 4, of the naturally occurring C-terminal amino acids are deleted or substituted.
  • [0052]
    Oligopeptides contained within the above sequence of particular interest include:
    (VIIIa) (49)
    Y-Arg-Val-Val-Gln-Arg-Glu-Lys-Arg-Z-X
    (VIIIb) (50)
    Y-Pro-Thr-Lys-Ala-Lys-Arg-Arg-Val-Val-
    Gln-Arg-Glu-Lys-Arg-X.
  • [0053]
    The next peptides of interest will be derived from the env region known as gp41.
  • [0054]
    The next peptide, IX (56), will be encoded by the region extending from about bp 7498 to bp 7554, where oligopeptides included within the following sequence will include linear epitopes within such sequence:
    (IX) (56)
    Ile-Lys-Gln-Leu-Gln-Ala-Arg-Ile-Leu-
    Ala-Val-Glu-Arg-Tyr-Leu-Lys-Asp-Gln-Gln-Z-X,
  • [0055]
    wherein X, Y and Z have been defined previously.
  • [0056]
    Oligopeptides contained within the above sequence of particular interest include:
    (IXa) (56/39)
    Y-Arg-Ile-Leu-Ala-Val-Glu-Arg-Tyr-Leu-
    Lys-Asp-Gln-Gln-Z-X
    and
    (IXb) (39/56)
    Y-Ile-Lys-Gln-Leu-Gln-Ala-Arg-Ile-Leu-Z-X.
  • [0057]
    The next peptide of interest, X (39), will be encoded by the region from about bp 7516 through bp 7593 and has the following amino acid sequence, where oligopeptides included within the following sequence will include linear epitopes within such sequence:
    (X) (39)
    Arg-Ile-Leu-Ala-Val-Glu-Arg-Tyr-Leu-Lys-
    Asp-Gln-Gln-Leu-Leu-Gly-Ile-Trp-Gly-Cys-
    Ser-Gly-Lys-Leu-Ile-Cys-X, where X is OH or NH2.
  • [0058]
    The next peptide, XI (40), will be encoded by the region extending from about bp 7630 to 7689 where oligopeptides included within the following sequence will include linear epitopes within such sequence:
    (XI) (40)
    Y-Lys-Ser-Leu-Glu-Gln-Ile-Trp-Asn-Asn-
    Met-Thr-Trp-Met-Glu-Trp-Asp-Arg-Glu-
    Ile-Asn-Z-X,
  • [0059]
    wherein Y, X and Z have been defined previously.
  • [0060]
    The next peptide of interest, XII (23), will be encoded by the region extending from about bp 7708 through bp 7779. This peptide will include any oligopeptides coding for linear epitopes within the following amino acid sequence:
    (XII) (23)
    Y-His-Ser-Leu-Ile-Glu-Glu-Ser-Gln-Asn-
    Gln-Gln-Glu-Lys-Asn-Glu-Gln-Glu-Leu-Leu-
    Glu-Leu-Asp-Lys-Trp-Z-X,
  • [0061]
    wherein X, Y and Z have been defined previously.
  • [0062]
    The next peptide of interest, XIII (79), will be encoded by the region extending from about bp 7543 through bp 7593. This peptide will include any oligopeptides coding for linear epitopes within the following amino acid sequence:
    (XIII) (79)
    Y-Lys-Asp-Gln-Gln-Leu-Leu-Gly-Ile-Trp-Gly-
    Cys-Ser-Gly-Lys-Leu-Ile-Cys-X,
  • [0063]
    wherein X and Y have been previously defined.
  • [0064]
    The next peptide of interest, XIIIa (78), will be encoded by the region extending from about bp 7561 through bp 7593. This peptide will include any oligopeptide coding for linear epitopes within the following amino acid sequence:
    (XIIIa) (78)
    Gly-Ile-Trp-Gly-Cys-Ser-Gly-Lys-Leu-Ile-Cys.
  • [0065]
    Of particular interest is the use of the mercaptan group of cysteines or thioglycolic acids used for acylating terminal amino groups or the like for linking two of the peptides or oligopeptides or combinations thereof by a disulfide linkage or a longer linkage. To achieve this, compounds may be employed having bis-haloacetyl groups, nitroarylhalides, or the like, where the reagents are specific for thio groups. Thus, the linking between the two mercapto groups of the different peptides or oligopeptides may be a single bond or a linking group of at least two, usually at least four, and not more than about 16, usually not more than about 14 carbon atoms. Of particular interest, is where a member of a sequence from the gag region is linked to a member from the env region. These chimeric peptides which may include non-amino acid linkages, may be further modified as will be described for the peptides and oligopeptides.
  • [0066]
    The subject peptides may be employed linked to a soluble macromolecular (e.g., ≧5 kDal) carrier. Conveniently, the carrier may be a poly(amino acid), either naturally occurring or synthetic, to which antibodies are unlikely to be encountered in human serum. Illustrative polypeptides include poly-L-lysine, bovine serum albumin, keyhole limpet hemocyanin, bovine gamma globulin, etc. The choice is primarily one of convenience and availability.
  • [0067]
    With such conjugates, there will be at least one molecule of at least one subject peptide per macromolecule and not more than about 1 per 0.5 kDal, usually not more than about 1 per 2 kDal of the macromolecule. One or more different peptides may be linked to the same macromolecule.
  • [0068]
    The manner of linking is conventional, employing such reagents as p-maleimidobenzoic acid, p-methyldithiobenzoic acid, maleic acid anhydride, succinic acid anhydride, glutaraldehyde, etc. The linkage may occur at the N-terminus, C-terminus or at a site intermediate the ends of the molecule. The subject peptide may be derivatized for linking, may be linked while bound to a support, or the like.
  • [0069]
    The compounds may be employed as labeled or unlabeled compounds depending upon their use. (By label is intended a molecule which provides, directly or indirectly, a detectable signal.) Various labels may be employed, such as radionuclides, enzymes, fluorescers, chemiluminescers, enzyme substrates, cofactors or inhibitors, particles, e.g., magnetic particles, combinations of ligands and receptors, e.g., biotin and avidin, or the like. In addition, the peptides may be modified in a variety of ways for binding to a surface, e.g., microtiter plate, glass beads, chromatographic surface, e.g., paper, cellulose, silica gel, or the like. The particular manner in which the polypeptides are joined to another compound or surface is conventional and finds ample illustration in the literature. See, for example, U.S. Pat. Nos. 4,371,515; 4,487,715; and patents cited therein.
  • [0070]
    Various assay protocols may be employed for detecting the presence of either antibodies to retroviral proteins or retroviral proteins themselves. Of particular interest is using the peptide as the labeled reagent, where the label allows for a detectable signal, or binding the peptide, either directly or indirectly to a surface, where antibody to the peptide in the sample will become bound to the peptide on the surface. The presence of human antibody bound to the peptide can then be detected by employing a xenogeneic antibody specific for human immunoglobulin, normally both human IgM and IgG, or a labeled protein specific for immune complexes, e.g., Rf factor or S. aureus protein A.
  • [0071]
    Various heterogeneous protocols may be employed, either competitive or non-competitive. Peptide may be bound to a surface or support (“support”) and labeled antibody allowed to compete with antibody in the sample for the limited amount of bound peptide. The amount of label bound to the support would be related to the amount of competitive antibody in the sample.
  • [0072]
    Antibody could be bound to the support and the sample combined with labeled peptide. After contact of the reaction mixture with the bound antibody, the amount of label bound to the support would relate to the amount of cognate antibody in the sample.
  • [0073]
    Xenogeneic anti-human antibody, e.g., antibodies to the Fc of IgG and IgM (immunoglobulins), could be bound to a support. The sample would be contacted with the immunoglobulins and labeled peptide, whereby the amount of labeled peptide bound to the support would be indicative of the presence of the cognate antibodies.
  • [0074]
    Alternatively, homogeneous assays can be employed where the peptide is bound to an enzyme, fluorescer, or other label, where the binding of antibody to the peptide results in being able to discriminate between the label involved with a specific binding pair complex and label which is not involved in the complex. For assays involving such techniques, see for example U.S. Pat. Nos. 3,817,837; 3,850,752; 3,901,654; 3,935,074; 3,984,533; 3,996,345; 4,034,074; and 4,098,876, whose disclosures are incorporated herein by reference.
  • [0075]
    As an illustration of the subject invention the subject peptides may be conjugated to a fluorescent molecule, such as fluorescein, rhodamine or umbelliferone. Various techniques may be used for detecting complex formation with antibodies, e.g., fluorescence polarization. In this assay the fluorescence polarization is different between complexed and uncomplexed peptide conjugate. Apparatuses are available for measuring changes in fluorescence polarization, e.g., TDx supplied by Abbott Laboratories, Chicago, Ill.
  • [0076]
    Illustrative of an assay technique is the use of sample containers, e.g., microtiter plate wells, where the subject polypeptides or conjugates thereof are adhered to the container bottom and/or walls either covalently or non-covalently. The sample, normally human blood or serum diluted in an appropriately buffered medium, is added to the container and a sufficient time allowed for complex formation between the polypeptide(s) and any cognate antibodies in the sample. The supernatant is removed and the container washed to remove non-specifically bound proteins.
  • [0077]
    A labeled specific binding protein which specifically binds to the complex is employed for detection. To the container may be added xenogeneic antisera to human immunoglobulin, particularly anti-(human IgM and IgG) in an appropriately buffered medium. The xenogeneic antisera will normally be labeled with a detectable label, e.g., radionuclide or enzyme. Instead of antisera, proteins specific for the immune complex may be employed, e.g., S. aureus protein A. The label may then be detected. For example, with an enzyme, after removal of non-specifically bound enzyme label, a developer solution is added. The developer solution will contain an enzyme substrate and possibly enzyme cofactors, chromogens, etc., which, upon reaction, provide a colored or fluorescent product which may be detected calorimetrically or fluorimetrically, respectively.
  • [0078]
    The peptides can be prepared in a wide variety of ways. The peptides, because of their relatively short size, may be synthesized in solution or on a solid support in accordance with conventional techniques. Various automatic synthesizers are commercially available today and can be used in accordance with known protocols. See, for example, Stewart and Young, Solid Phase Peptide Synthesis, 2nd ed., Pierce Chemical Co., 1984; and Tam et al., J. Am. Chem. Soc. (1983) 105:6442.
  • [0079]
    Alternatively, hybrid DNA technology may be employed where a synthetic gene may be prepared by employing single strands which code for the polypeptide or substantially complementary strands thereof, where the single strands overlap and can be brought together in an annealing medium so as to hybridize. The hybridized strands may then be ligated to form the complete gene and by choice of appropriate termini, the gene may be inserted into expression vectors, which are readily available today. See, for example, Maniatis et al., Molecular Cloning, A Laboratory Manual, CSH, Cold Spring Harbor Laboratory, 1982. Or, the region of the viral genome coding for the peptide may be cloned by conventional recombinant DNA techniques and expressed (see Maniatis, supra).
  • [0080]
    DNA coding sequences which may be used for expressing peptides I-XIII are:
    I (15) (TAT) GATTGTAAGACTATTTTAAAAGCATTGGGACCAG
    CAGCTACACTAGAAGAAATGATGACAGCATGT
    II (17) (TGT) TTAAAAGAGACCATCAATGAGGAAGCTGCAGAAT
    GGGATAGAGTGCATCCAGTGCATGCA
    III (92) GATAGAGTGCATCCAGTGCATGCAGGGCCTATTGCACCA
    GGCCAG
    IV (90) TATAGCCCTACCAGCATTCTGGACATAAGACAAGGACCA
    AAAGAACCCTTTAGAGACTATGTAGACCGGTTCTATAAA
    ACTCTAAGA
    V (88) AATTGGATGACAGAAACCTTGTTGGTCCAAAATGCGAAC
    CCAGATTGTAAGACTATTTTAAAAGCATTGGGACCAGCA
    GCTACACTAGAAGAAATGATGACAGCATGT
    VI (97) AGGGAGCTAGAACGATTCGCTGTTAATCCTGGCCTGTTA
    GAAACATCAGAAGGCTGTAGACAAATACTGGGACAGCTA
    CAACCATCCCTTCAGACA
    VII (71) GACACAGGACACAGCAGCCAGGTCAGCCAAAATTAC
    VIII (36) GTAAAAATTGAACCATTAGGAGTAGCACCCACCAAGGCA
    AAGAGAAGAGTGGTGCAGAGAGAAAAAAGAGCA (TGT)
    IX (56) ATCAAGCAGCTCCAGGCAAGAATCCTGGCTGTGGAAAGA
    TACCTAAAGGATCAACAG (TGT)
    X (39) AGAATCCTGGCTGTGGAAAGATACCTAAAGGATCAACAG
    CTCCTGGGGATTTGGGGTTGCTCTGGAAAACTCATTTGC
    XI (40) (TGT) AAATCTCTGGAACAGATTTGGAATAACATGACCT
    GGATGGAGTGGGACAGAGAAATTAAC (TGT)
    XII (23) (TGT) CATTCCTTAATTGAAGAATCGCAAAACCAGCAAG
    AAAAGAATGAACAAGAATTATTGGAATTAGATAAATGG
    (GGA)
    XIII (79) AAGGATCAACAGCTCCTGGGGATTTGGGGTTGCTCTGGA
    AAACTCATTTGC
  • [0081]
    Fragments from these sequences may be employed for expression of peptide fragments, conservative base changes can be made, where the modified codon(s) code for the same amino acid(s), or non-conservative changes in the coding sequence may be made, where the resulting amino acid may be a conservative or non-conservative change.
  • [0082]
    The coding sequence may be extended at either the 5′- or 3′-terminus or both termini to extend the peptide, while retaining its epitopic site. The extension may provide for an arm for linking, e.g., to a label, such as an enzyme, for joining two or all of the peptides together in the same chain, for providing antigenic activity, or the like.
  • [0083]
    For expression, the coding sequence will be provided with start and stop codons, promoter and terminator regions and usually a replication system to provide an expression vector for expression in a cellular host, e.g., prokaryotic or eukaryotic, bacterial, yeast, mammal, etc.
  • [0084]
    The sequences by themselves, fragments thereof, or larger sequences, usually at least 15 bases, preferably at least 18 bases, may be used as probes for detection of retroviral RNA or proviral DNA. Numerous techniques are described, such as the Grunstein-Hogness technique, Southern technique, Northern technique, dot-blot, improvements thereon, as well as other methodology. See, for example, WO 83/02277 and Berent et al., Biotechniques (1985) 3:208.
  • [0085]
    Conveniently, the polypeptides may be prepared as fused proteins, where the polypeptide may be the N- or C-terminus of the fused polypeptide. The resulting fused protein could be used directly by itself as the reagent or the subject polypeptide may be cleaved from all or a portion of the remaining sequence of the fused protein. With a polypeptide where there are no internal methionines, by introducing a methionine at the fusion site, the polypeptide may be cleaved employing cyanogen bromide. Where there is an internal methionine, it would be necessary to provide for a proteolytic cleavage site, e.g., poly-lysine and/or -arginine or combinations thereof, or the internal methionine could be substituted with an amino acid such as leucine and an N-terminal methionine added for cyanogen bromide cleavage. A wide variety of proteases, including dipeptidases, are well known and the appropriate processing signal could be introduced at the proper site. The processing signal may have tandem repeats so as to insure cleavage, since the presence of one or more extraneous amino acids will not interfere with the utility of the subject polypeptides.
  • [0086]
    Depending upon the nature of the assay, the physiological sample, e.g., saliva, blood, plasma, or serum, may be pretreated by dilution into an assay medium, which will usually be an aqueous buffered medium employing one of a variety of buffers, such as phosphate, tris, or the like. A preferred diluent is blotto (5% w/v nonfat dry milk, 0.01% thimerosal, 0.01% Antifoam A in 0.01 M sodium phosphate, pH 7.2, and 0.15 M NaCl). Usually the pH will be in the range of about 6 to 9. The sample will then be combined with the reagent in accordance with the appropriate protocol and sufficient time allowed for binding. Where a heterogeneous system is used, usually the stages will be followed by washes, to minimize non-specific binding. At the end of the procedure, the label will be detected in accordance with conventional ways.
  • [0087]
    Besides the use of the subject peptides and their analogs in assays, the subject peptides may also find use by themselves or in combination in vaccines. The peptides may be formulated in a convenient manner, generally at concentrations in the range of 1 μg to 20 mg/kg of host. Physiologically acceptable media may be used as carriers, such as sterile water, saline, phosphate buffered saline, and the like. Adjuvants may be employed, such as aluminum hydroxide gel, or the like. Administration may be by injection, e.g., intramuscularly, peritoneally, subcutaneously, intravenously, etc. Administration may be one or a plurality of times, usually at one to four week intervals.
  • [0088]
    The following examples are offered by way of illustration and not by way of limitation.
  • EXPERIMENTAL
  • [0089]
    Peptides 15, 71, 88, 90, 92 and 97 were assembled on a t-butyloxycarbonyl (BOC)-methylbenzyl-cysteine-phenyl-acetamidomethyl (PAM) polystyrene/divinylbenzene resin (Applied Biosystems, Inc., Foster City, Calif.). For carboxamide peptides 78 and 79 p-methylbenzhydrylamine polystyrene/divinylbenzene was used. Symmetrical anhydride couplings were carried out in an Applied Biosystems 430A synthesizer, except that glutamine and asparagine were coupled as hydroxybenzotriazole esters. Benzyl based side chain protection and BOC alpha-amine protection were used. Tryptophan was protected by the formyl moiety, methionine was protected by its sulfoxide, and dinitrophenol was used for protecting histidine. Protecting groups were removed by conventional procedures.
  • [0090]
    Peptide 36 was assembled on a benzhydrylamine polystyrene/divinylbenzene resin in a Beckman 990 peptide synthesizer (Beckman Instruments, La Brea, Calif.). Benzyl based side chain protection and BOC alpha-amine protection were used. All the residues were added by the direct dicyclohexylcarbodiimide method, except for glutamine which was coupled as the hydroxybenzotriazole ester.
  • [0091]
    Peptide 39 was synthesized on a benzhydrylamine resin as described for peptide 36 with asparagine also being coupled as the ester.
  • [0092]
    When the peptides were radiolabeled, it was by acetylating the amino terminus with 3H-acetic acid and an excess of dicyclohexylcarbodiimide.
  • [0093]
    The peptides were deprotected and cleaved from the resin by the Tam “low-high” HF protocol (Tam et al., supra). Peptides 36, 39, 79, 78, 88, 90, 92 and 97 were extracted from the resin in 5% acetic acid and subjected to gel filtration chromatography in 5% acetic acid. Peptides 15 and 71 were extracted in 0.5M ammonium carbonate/0.001M dithiothreitol (DTT) and chromatographed in 0.05M ammonium carbonate/0.005M β-mercaptoethanol. Fractions containing the peptide were pooled and lyophilized. The integrity of the synthetic products was assured by ninhydrin monitoring after each coupling and by analytical reverse phase chromatography and amino acid analysis.
  • [0094]
    Peptides 90, 92 and 97 were polymerized by oxidation of their sulfhydryls to intermolecular disulfides. Briefly, the lyophilized reduced peptide was dissolved in minimal 6M guanidine HCl/0.1M sodium phosphate, pH 9.0, and allowed to oxidize overnight at room temperature.
  • [0095]
    Peptides 15, 23, 36, 40, 49, 50 and 56 synthesized above were conjugated to bovine serum albumin (BSA) which had been derivatized with N-succinimidyl-4-(N-maleimidomethyl)-cyclohexane-1-carboxylate (SMCC), essentially as described by Ishikawa et al., J. of Immunoassay (1983) 4:209.
  • [0096]
    To 2 ml of a BSA solution (20 mg/ml in 0.1M potassium phosphate, pH 7.0) at 30° C. was added 1.5 ml of an SMCC solution (8 mg/ml in dimethylformamide). The mixture was stirred magnetically for 1 hr, after which it was centrifuged to remove any precipitated albumin. The clarified mixture was then subjected to gel filtration on Sephadex G-25 equilibrated in 0.1M potassium phosphate, pH 6.0. The protein-containing fractions, as determined by their absorbance at 280 nm, were pooled and stored frozen at −70° C. until needed.
  • [0097]
    The peptides synthesized above were dissolved in 0.1M sodium phosphate, pH 8.0 to a concentration of 5 mg/ml (peptide 36), 8 mg/ml (peptide 15) or 1.6 mg/ml (peptide 39). To 1.5 ml of each solution was added 2 mg of solid DTT. The solutions were stirred for 30 min at 30° C., after which they were subjected to gel filtration chromatography on Sephadex G-10, equilibrated in 0.1M potassium phosphate, pH 6.0. The tritium-containing fractions, as determined by scintillography of aliquots, were pooled and mixed with 1 ml (0.5 ml for peptide V) of SMCC-derivatized BSA. The resultant mixtures were stirred at 30° C. for 12 hr and then dialyzed exhaustively against water.
  • [0098]
    The other peptides were prepared in accordance with the procedures described above and conjugated to BSA in accordance with the above described procedures. The ratio of peptide to BSA was determined by employing radiotracers in accordance with conventional ways.
    mols peptide
    mol BSA
    I (15) 14
    II (17)  5
    VIII (36)  9
    IX (56) 17
    X (39)  6
    XI (40) 18
    XII (23)  30*
  • [0099]
    Analysis by ELISA
  • [0100]
    The lyophilized peptide or protein/peptide conjugate was dissolved in 6M guanidine HCl. The guanidine solutions were diluted in 0.05M carbonate/bicarbonate buffer (pH 9.6) to a final peptide concentration of 8 to 40 μg/ml just prior to plating in the 96-well plates. Fifty μl of peptide solution were aliquoted per microtiter well and incubated at 4° C. overnight. Plates were then blocked with BLOTTO (5% [w/v] nonfat dry milk/0.01% thimerosal/0.01% antifoam A in 0.01M sodium phosphate, pH 7.2/0.15M sodium chloride) for one hour at 37° C. Sera were diluted 1:100 with a 1:1 mixture of BLOTTO and PBS (0.01M sodium phosphate, pH 7.3/0.15M NaCl), and 50 μl of diluted sera was added to each well and incubated for one hour at 37° C. The sera were removed and the plates were washed three times in wash buffer (0.15M NaCl/0.05% [w/v] Tween 20) before adding 100 μl of the goat anti-human IgG/horseradish peroxidase conjugate (50% stock diluted 1:10,000 in 50 mM sodium citrate/0.05% Tween 20/1% heat-inactivated normal goat serum; obtained from Antibodies, Inc., Davis, Calif.) for one hour at 37° C. The conjugate was removed and the plates washed three times with 0.15M NaCl/0.05% (w/v) Tween 20. The ELISA was developed by adding 100 μl per well of substrate solution (10 mg 3,3′, 5,5′-tetramethylbenzidine in 50 ml 0.05M sodium citrate, pH 7.0) for 30 min at room temperature. Reactions were stopped with 100 μl per well of 3N H2SO4, and the optical density at 450 nm determined by an automated ELISA reader.
  • [0101]
    Summary of Table 1
  • [0102]
    Table 1 gives ELISA results for all petpides that are immunoreactive.
  • [0103]
    Peptides 49 and 50 are part of peptide 36.
  • [0104]
    Peptide 56 partially overlaps peptide 39.
  • [0105]
    Peptide 49-BSA reactive with {fraction (10/10)} positve sera; not reactive with {fraction (2/2)} negative sera.
  • [0106]
    Peptide 50-BSA reactive with {fraction (10/10)} positive sera; not reactive with {fraction (2/2)} negative sera.
  • [0107]
    Peptide 56-BSA reactive with {fraction (10/10)} positive sera; not reactive with {fraction (2/2)} negative sera.
  • [0108]
    Peptide 40-BSA reactive with {fraction (10/10)} positive sera; not reactive with {fraction (2/2)} negative sera.
  • [0109]
    Peptide 23-BSA reactive with {fraction (10/10)} positive sera; not reactive with {fraction (2/2)} negative sera.
  • [0110]
    Peptide 15-BSA reactive with {fraction (10/10)} positive sera; not reactive with {fraction (2/2)} negative sera.
  • [0111]
    Peptide 36-BSA reactive with {fraction (9/10)} positive sera; not reactive with {fraction (2/2)} negative sera.
  • [0112]
    In a larger panel, peptide 56 which partially overlaps peptide 39, is not reactive with all sera that are reactive with peptide 39. This suggests that there are at least two reactive epitopes within peptide 39 or that peptides 39 and 56 contain non overlapping reactive epitopes.
  • [0113]
    Peptide 23 (both conjugated to BSA and unconjugated) was further tested against a larger panel of sera (23 positives, 8 negatives) and displays a sensitivity of 80-90%.
  • [0114]
    Summary of Table 2
  • [0115]
    Table 2 shows that two of the peptides derived from the gag region (#15 and #17) are reactive with LAV seropositive sera that are poorly reactive or unreactive with peptide 39. This supports the use of a combination of gag and env peptides to produce a more sensitive assay.
  • [0116]
    Summary of Table 3
  • [0117]
    Table 3 compares results-obtained with peptides 15-BSA and 39 with results obtained with these peptides physically mixed (15-BSA+39) or chemically combined (thiol-oxidized 15+39).
  • [0118]
    The result obtained when positive samples are assayed with either the physical or chemical combination of peptides 15 and 39 is generally higher than that obtained with either peptide alone. This is clearly demonstrated with samples 126, 131, 135, 138 and 1296.
  • [0119]
    Summary of Table 4
  • [0120]
    Table 4 compares results obtained with peptides 71, 78, 79, 88, 90, 92 and 97 in an ELISA assay. All of the peptides except one provide better than 70% correlation for positives and two peptides had 100% correlation.
    TABLE 1
    COMPARISON OF PEPTIDES WITH A WHOLE VIRUS LYSATE IN
    AN ELISA ASSAY FOR THE DETECTION OF ANTIBODIES TO LAV
    ELISA Using
    Positive Whole Virus Confirmed as BSA- BSA- BSA- BSA- BSA- BSA- BSA-
    Sera Diagnosis Lysate1 Serpositive2 Pep15 Pep36 Pep49 Pep50 Pep39 Pep56 Pep40 Pep23 Pep 23
    155 LAS and/or 1.069 yes 0.525 1.679 0.955 1.678 1.167 1.675 0.603 1.640 0.111
    homosexual
    124 LAS and/or 1.189 yes 1.329 1.465 1.334 2.207 1.073 1.842 1.462 2.117 2.127
    homosexual
    138 LAS and/or 1.302 yes 0.378 0.159 0.204 0.374 0.514 0.643 0.774 0.960 0.106
    homosexual
    133 LAS and/or 1.250 yes 0.365 0.567 0.409 0.581 1.036 0.627 1.297 2.077 N.D.
    homosexual
    131 LAS and/or 1.220 yes 0.411 0.272 0.225 0.595 0.448 1.679 1.209 1.621 0.949
    homosexual
    134 LAS and/or 1.050 yes 0.559 0.712 0.729 0.293 1.619 2.170 0.567 1.705 1.552
    homosexual
    153 LAS and/or 2.000 yes 0.467 0.548 1.011 0.591 1.314 1.324 0.734 0.970 0.524
    homosexual
    157 LAS and/or 1.349 yes 0.366 0.321 0.148 0.427 1.326 2.179 1.153 2.017 1.158
    homosexual
    Y-1/ LAS and/or 2.000 yes 2.109 1.022 1.547 1.928 1.305 2.115 1.257 1.565 0.762
    CDC homosexual
    501 LAS and/or 1.109 yes 2.374 1.168 1.938 2.209 1.167 1.170 0.625 0.467 0.059
    homosexual
    1892  Healthy n.d. n.d.4 0.128 0.113 0.119 0.124 0.045 0.143 0.141 0.253 0.034
    heterosexual
    639 Healthy 0.123 not 0.159 0.142 0.102 0.186 0.038 0.355 0.251 0.286 0.024
    heterosexual seropositive
  • [0121]
    [0121]
    TABLE 2
    COMPARISON OF GAG PEPTIDES WITH PEPTIDE 39 IN AN
    ELISA ASSAY FOR THE DETECTION OF ANTIBODIES TO LAV
    Serum Whole Virus Confirmed as
    No. Diagnosis Lysate1 Seropositive2 17-BSA 15-BSA 39
    1296 Blood Donor 2.01 yes 0.633 0.65 0.11
    501 Unknown 1.109 yes 0.18 2.04 2.15
    129 LAS and/or 1.08 yes 0.62 0.49 0.42
    homosexual
    154 LAS and/or 1.41 yes 0.26 0.26 0.35
    homosexual
    7 LAS and/or 2.00 yes 0.79 1.02 0.22
    homosexual
    641 Healthy 0.20 n.d. 0.22 0.19 0.04
    heterosexual
    639 Healthy 0.12 n.d. 0.20 0.16 0.05
    heterosexual
  • [0122]
    [0122]
    TABLE 3
    COMPARISON OF PEPTIDES 15 and 39 INDIVIDUALLY WITH PEPTIDES 15 AND 39
    PHYSICALLY OR CHEMICALLY COMBINED IN AN ELISA ASSAY
    FOR THE DETECTION OF ANTIBODIES TO LAV
    Serum Whole Virus Confirmed as 15-BSA Thiol-oxidized
    No. Diagnosis Lysate1 Seropositive2 15-BSA 39 39 15 + 39
    133 LAS and/or 1.250 yes 0.13 1.02 >2 1.88
    homosexual
    134 LAS and/or 1.050 yes 0.21 1.62 >2 2.27
    homosexual
    135 LAS and/or 1.310 yes 0.25 0.32 1.93 1.48
    homosexual
    138 LAS and/or 1.302 yes 0.13 0.51 1.65 0.91
    homosexual
    153 LAS and/or 2.000 yes 0.16 1.32 n.d. 1.89
    homosexual
    154 LAS and/or 1.41 yes 0.19 0.35 n.d. 1.35
    homosexual
    155 LAS and/or 1.069 yes 0.29 1.17 n.d. 1.83
    homosexual
    157 LAS and/or 1.349 yes 0.14 1.33 n.d. n.d.
    homosexual
    666 Unknown 2.000 yes 1.60 1.39 >2 2.01
    1296  Blood Donor 2.00 yes 0.65 0.11 0.99 0.16
    633 Healthy 0.222 not sero- 0.09 0.05 n.d. n.d.
    heterosexual positive
    637 Healthy 0.097 not sero- 0.13 0.04 0.42 n.d.
    heterosexual positive
    639 Healthy 0.123 not sero- 0.12 0.04 0.22 0.11
    heterosexual positive
    641 Healthy 0.199 not sero- 0.18 0.03 0.49 0.13
    heterosexual positive
    501 Positive 1.109 yes 1.39 1.17 >2.0 1.77
    control
    Y-1 Positive 2.000 yes 1.02 1.30 >2.0 2.02
    CDC control pool
    120 LAS3 and/or 1.540 yes 0.19 1.37 n.d. n.d.
    homosexual
    121 LAS and/or 1.483 yes 0.09 1.51 >2.0 1.96
    homosexual
    122 LAS and/or 1.283 yes 0.14 1.88 >2.0 2.33
    homosexual
    124 LAS and/or 1.189 yes 0.60 1.06 n.d. n.d.
    homosexual
    125 LAS and/or 1.232 yes 0.18 1.53 n.d. n.d.
    homosexual
    126 LAS and/or 1.233 yes 0.24 0.51 >2 1.5
    homosexual
    127 LAS and/or 1.046 yes 0.25 1.52 n.d. n.d.
    homosexual
    128 LAS and/or 1.284 yes 0.09 1.07 n.d. n.d.
    homosexual
    129 LAS and/or 1.081 yes 0.33 0.42 n.d. n.d.
    homosexual
    130 LAS and/or 0.912 yes 0.28 1.17 n.d. n.d.
    homosexual
    131 LAS and/or 1.220 yes 0.14 0.45 >2 1.22
    homosexual
    132 LAS and/or 1.237 yes 0.15 1.24 >2 1.91
    homosexual
    667 Healthy 0.095 n.d. 0.15 0.04 0.42 n.d.
    heterosexual
    1890  Healthy n.d. n.d. 0.15 0.05 0.39 0.15
    heterosexual
    1891  Healthy n.d. n.d. 0.17 0.05 0.31 0.12
    heterosexual
    1892  Healthy n.d. n.d. 0.08 0.05 0.18 0.07
    heterosexual
  • [0123]
    [0123]
    TABLE 4
    COMPARISON OF PEPTIDES 92, 90, 88, 97, 71, 79 and 78
    IN AN ELISA ASSAY FOR THE DETECTION OF
    ANTIBODIES TO LAV
    Confirmed
    Serum as
    No. Seropositive 92 90 88 97 71 79 78
    127 yes .201 1.256 1.610 2.558 .476 2.346 .520
    130 yes .220 .900 1.912 2.341 .350 1.808 .438
    124 yes .105 1.175 .372 2.302 .514 1.086 .092
    125 yes .126 1.386 1.798 .395 .416 2.266 .281
    128 yes .122 .882 .201 .377 .246 1.144 .123
    134 yes .131 1.159 .358 2.455 .535 1.316 .118
    135 yes .120 .644 .157 1.231 .292 .381 .119
    153 yes .138 1.150 .180 .780 .352 1.039 .146
    154 yes ND .623 .256 .365 .210 ND ND
    155 yes .108 .845 .058 1.984 .185 1.584 .105
    157 yes .118 .936 .942 1.620 .536 1.162 .146
    120 yes .159 1.031 .740 .221 .362 1.546 .239
    121 yes .157 1.284 1.776 .396 .307 2.084 .205
    132 yes .100 .909 .422 .399 .398 1.386 .192
    138 yes .086 .495 ND 1.201 .285 .312 .093
    133 yes .100 .739 .143 .526 .312 .597 .114
    131 yes .112 .841 .197 .742 .188 1.150 .101
    501 yes .472 1.098 2.058 2.253 .341 1.768 .216
    129 yes .091 ND ND ND ND .562 .085
    Y1 yes ND ND 2.228 ND ND ND ND
    N3 no .074 .603 .106 .162 .101 .224 .076
    N12 no .075 .617 .131 .174 .088 .174 .056
    N4 no .058 .519 .128 .190 .090 .172 .040
    639 no .082 .474 .092 .115 .121 .153 .059
    641 no .081 .369 .090 .155 .169 .140 .085
    N13 no .079 .455 .111 .120 .100 .226 .122
    N14 no .054 .560 .098 .151 .085 .162 .070
    N16 no .077 .521 .083 .122 .070 .183 .079
    Cutoff 0.10 0.70 0.20 0.20 0.20 0.30 0.20
    Fraction of Confirmed 14/18 15/18 13/18 18/18 16/18 18/18 6/18
    Seropositive Samples
    Detected as Positive
  • FOOTNOTES TO TABLES 1-3
  • [0124]
    [0124]1Prepared as described in U.K. application Serial No. 83/24800, filed Sep. 15, 1983.
  • [0125]
    [0125]2 Radiolabeled LAV antigens were disrupted in RIPA buffer (Gilead et al., Nature (1976) 264:263) and then were reacted with human serum. The resultant immune complexes were separated by binding to a Staphylococcus aureus adsorbent (Kessler, J. Immunology (1975) 115:1617) followed by multiple washings. Immuneprecipitated antigens were analyzed by SDS polyacrylamide gel electrophoresis (Laemmli, Nature (1970) 227:680) followed by fluorography. Presence of either a p25 or gp43 band was considered necessary and sufficient to confirm a sample as seropositive.
  • [0126]
    [0126]3 LAS=lymphadenopathy syndrome.
  • [0127]
    [0127]4 N.D.=not determined.
  • [0128]
    It is evident from the foregoing results that by employing one or a combination of peptides of the subject invention, a sensitive accurate test for the presence of antibodies to AIDS is provided. The subject peptides can be used by themselves or in combination with a screening assay or confirmatory assay, where the complete lysate or complete antigens may be employed as an independent procedure. Furthermore, because of the specificities of the peptides, one would anticipate that the DNA sequences coding for the peptides would also find similar specificity in a DNA hybridization assay. Thus, the subject invention allows for the detection of patients who have been exposed to the retroviral etiologic agent of lymphadenopathy syndrome and/or AIDS.
  • [0129]
    Although the foregoing invention has been described in some detail by way of illustration and example for purposes of clarity of understanding, it will be obvious that certain changes and modifications may be practiced within the scope of the appended claims.

Claims (36)

What is claimed is:
1. In a method for detecting the presence of LAV/HTLV-III virus or antibody to LAV/HTLV-III virus where a sample is combined with a composition having epitopic sites immunologically competitive with LAV/HTLV-III epitopic sites, whereby antibodies bind to such protein composition to form a specific binding pair complex and the amount of complex formation is determined, the improvement which comprises:
employing in the assay medium as a reagent a compostion, containing at least one peptide which has at least five amino acids and fewer than 50 amino acids and is encoded for by the coding region of LAV/HTLV-III from bp 900 to bp 1421 or bp 7210 to bp 7815.
2. A method according to claim 1, wherein said coding region is from bp 1320 to bp 1388 or bp 7231 to bp 7794.
3. A method according to claim 2, wherein said composition includes at least two peptides, one encoded in the region bp 1320 to bp 1385 and the other encoded in the region bp 7231 to bp 7794.
4. A method according to claim 3, where each determination is made individually for the binding to each of the peptides.
5. In a method for detecting the presence of LAV/HTLV-III virus or antibody to LAV/HTLV-III virus where a sample is combined with a composition having epitopic sites immunologically competitive with LAV/HTLV-III epitopic sites, whereby antibodies bind to each such protein composition to form at least one specific binding pair complex and the amount of complex formation is determined, the improvement which comprises:
employing in the assay medium as a reagent a composition, containing at least one peptide which has at least five amino acids in a sequence which comes within the sequence of at least one of the following peptide sequences:
(I) (15) Y-Asp-Cys-Lys-Thr-Ile-Leu-Lys-Ala-Leu- Gly-Pro-Ala-Ala-Thr-Leu-Glu-Glu-Met-Met- Thr-Ala-Cys-X (II) (17) Y-Leu-Lys-Glu-Thr-Ile-Asn-Glu-Glu-Ala- Ala-Glu-Trp-Asp-Arg-Val-His-Pro-Val-His- Ala-Z-X (III) (92) Y-Asp-Arg-Val-His-Pro-Val-His-Ala-Gly-Pro-Ile-Ala-Pro- Gly-Gln-X (IV) (90) Y-Tyr-Ser-Pro-Thr-Ser-Ile-Leu-Asp-Ile-Arg- Gln-Gly-Pro-Lys-Glu-Pro-Phe-Arg-Asp-Tyr-Val- Asp-Arg-Phe-Tyr-Lys-Thr-Leu-Arg-Z-X (V) (88) Y-Asn-Trp-Nor-Thr-Glu-Thr-Leu-Leu-Val-Gln- Asn-Ala-Asn-Pro-Asp-Cys-Lys-Thr-Ile-Leu-Lys- Ala-Leu-Gly-Pro-Ala-Ala-Thr-Leu-Glu-Glu-Nor- Nor-Thr-Ala-Cys-X (VI) (97) Y-Arg-Glu-Leu-Glu-Arg-Phe-Ala-Val-Asn-Pro-Gly- Leu-Leu-Glu-Thr-Ser-Glu-Gly-Cys-Arg-Gln-Ile- Leu-Gly-Gln-Leu-Gln-Pro-Ser-Leu-Gln-Thr-X (VII) (71) Y-Asp-Thr-Gly-His-Ser-Ser-Gln-Val-Ser-Gln- Asn-Tyr (VIII) (36) Val-Lys-Ile-Glu-Pro-Leu-Gly-Val-Ala-Pro- Thr-Lys-Ala-Lys-Arg-Arg-Val-Val-Gln-Arg- Glu-Lys-Arg-Ala-Z-X (IX) (56) Ile-Lys-Gln-Leu-Gln-Ala-Arg-Ile-Leu- Ala-Val-Glu-Arg-Tyr-Leu-Lys-Asp-Gln-Gln-Z-X (X) (39) Arg-Ile-Leu-Ala-Val-Glu-Arg-Tyr-Leu-Lys- Asp-Gln-Gln-Leu-Leu-Gly-Ile-Trp-Gly-Cys- Ser-Gly-Lys-Leu-Ile-Cys-X (XI) (40) Y-Lys-Ser-Leu-Glu-Gln-Ile-Trp-Asn-Asn- Met-Thr-Trp-Met-Glu-Trp-Asp-Arg-Glu- Ile-Asn-Z-X (XII) (23) Y-His-Ser-Leu-Ile-Glu-Glu-Ser-Gln-Asn- Gln-Gln-Glu-Lys-Asn-Glu-Gln-Glu-Leu-Leu- Glu-Leu-Asp-Lys-Trp-Z-X (XIII) (79) Y-Lys-Asp-Gln-Gln-Leu-Leu-Gly-Ile-Trp-Gly- Cys-Ser-Gly-Lys-Leu-Ile-Cys-X,
where X is OH or NH2, and Y and Z, when present, are amino acids added to facilitate coupling, and where said peptide is free of other peptides or conjugated to a macromolecule for which antibodies in human sera are substantially absent.
6. A method according to claim 5, wherein said composition includes at least two of said peptides, at least one of said peptides having the amino acid sequence according to peptide sequences I to VII and at least one of said peptides having an amino acid sequence according to peptides VIII to XIII.
7. A method according to claim 6, wherein one of the peptides has one of the following sequences:
(VIIIa) (49) Y-Arg-Val-Val-Gln-Arg-Glu-Lys-Arg-X (VIIIb) (50) Y-Pro-Thr-Lys-Ala-Lys-Arg-Arg-Val-Val- Gln-Arg-Glu-Lys-Arg-X (IXa) (56/39) Y-Arg-Ile-Leu-Ala-Val-Glu-Arg-Tyr-Leu- Lys-Asp-Gln-Gln-Z-X (IXb) (39/56) Y-Ile-Lys-Gln-Leu-Gln-Ala-Arg-Ile-Leu-Z-X (XIIIa) (78) Gly-Ile-Trp-Gly-Cys-Ser-Gly-Lys-Leu-Ile-Cys,
where X is OH or NH2, and Y, when present, is an amino acid added to facilitate coupling.
8. A method according to claim 1, wherein said composition includes at least one peptide selected from p25 and one from p18.
9. A method according to claim 5, wherein said composition is bound to a solid surface.
10. A method for determining the presence of antibodies to LAV/HTLV-III in a physiological fluid, said method comprising:
introducing a human serum, plasma or blood sample into a sample container coated at least in part with at least one peptide having at least five amino acids which comes within the sequence of at least one of the following peptide sequences:
(I) (15) Y-Asp-Cys-Lys-Thr-Ile-Leu-Lys-Ala-Leu- Gly-Pro-Ala-Ala-Thr-Leu-Glu-Glu-Met-Met- Thr-Ala-Cys-X (II) (17) Y-Leu-Lys-Glu-Thr-Ile-Asn-Glu-Glu-Ala- Ala-Glu-Trp-Asp-Arg-Val-His-Pro-Val-His- Ala-X (III) (92) Y-Asp-Arg-Val-His-Pro-Val-His-Ala-Gly-Pro Ile-Ala-Pro-Gly-Gln-X (IV) (90) Y-Tyr-Ser-Pro-Thr-Ser-Ile-Leu-Asp-Ile-Arg Gln-Gly-Pro-Lys-Glu-Pro-Phe-Arg-Asp-Tyr-Val- Asp-Arg-Phe-Tyr-Lys-Thr-Leu-Arg-Z-X (V) (88) y-Asn-Trp-Nor-Thr-Glu-Thr-Leu-Leu-Val-Gln- Asn-Ala-Asn-Pro-Asp-Cys-Lys-Thr-Ile-Leu-Lys- Ala-Leu-Gly-Pro-Ala-Ala-Thr-Leu-Glu-Glu-Nor- Nor-Thr-Ala-Cys-X (VI) (97) Y-Arg-Glu-Leu-Glu-Arg-Phe-Ala-Val-Asn-Pro-Gly- Leu-Leu-Glu-Thr-Ser-Glu-Gly-Cys-Arg-Gln-Ile- Leu-Gly-Gln-Leu-Gln-Pro-Ser-Leu-Gln-Thr-X (VII) (71) Y-Asp-Thr-Gly-His-Ser-Ser-Gln-Val-Ser-Gln- Asn-Tyr (VIII) (36) Val-Lys-Ile-Glu-Pro-Leu-Gly-Val-Ala-Pro Thr-Lys-Ala-Lys-Arg-Arg-Val-Val-Gln-Arg Glu-Lys-Arg-Ala-Z-X (IX) (56) Ile-Lys-Gln-Leu-Gln-Ala-Arg-Ile-Leu- Ala-Val-Glu-Arg-Tyr-Leu-Lys-Asp-Gln-Gln-Z-X (X) (39) Arg-Ile-Leu-Ala-Val-Glu-Arg-Tyr-Leu-Lys Asp-Gln-Gln-Leu-Leu-Gly-Ile-Trp-Gly-Cys- Ser-Gly-Lys-Leu-Ile-Cys-X (XI) (40) Y-Lys-Ser-Leu-Glu-Gln-Ile-Trp-Asn-Asn- Met-Thr-Trp-Met-Glu-Trp-Asp-Arg-Glu- Ile-Asn-Z-X (XII) (23) Y-His-Ser-Leu-Ile-Glu-Glu-Ser-Gln-Asn- Gln-Gln-Glu-Lys-Asn-Glu-Gln-Glu-Leu-Leu- Glu-Leu-Asp-Lys-Trp-Z-X (XIII) (79) Y-Lys-Asp-Gln-Gln-Leu-Leu-Gly-Ile-Trp-Gly- Cys-Ser-Gly-Lys-Leu-Ile-Cys-X,
where X is OH or NH2, and Y and Z, if present, are amino acids added to facilitate coupling, wherein said peptides are free of other peptides or conjugated to a macromolecule for which antibodies in human sera are substantially absent;
incubating for a sufficient time for complex formation to occur; and
determining the formation of complex by employing a labeled specific binding protein which binds to said complex and provides a detectable signal.
11. A method according to claim 10, wherein said peptide is conjugated to a water soluble protein of at least 5 kDal as said macromolecule.
12. A method according to claim 10, wherein two of said peptides are covalently linked together through a bond or chain.
13. A method according to claim 10, wherein said specific binding protein is antibody to human immunoglobulin.
14. A method according to claim 10, wherein said composition includes at least two of said peptides, at least one peptide coming from peptides I to VII and at least one peptide coming from peptides VIII to XIII.
15. A method for determining the presence of antibodies to LAV/HTLV-III in a physiological fluid, said method comprising:
combining a human serum, plasma or blood sample with at least one labeled peptide having at least five amino acids in a sequence which comes within the sequence of at least one of the following peptide sequences:
(I) (15) Y-Asp-Cys-Lys-Thr-Ile-Leu-Lys-Ala-Leu- Gly-Pro-Ala-Ala-Thr-Leu-Glu-Glu-Met-Met Thr-Ala-Cys-X (II) (17) Y-Leu-Lys-Glu-Thr-Ile-Asn-Glu-Glu-Ala- Ala-Glu-Trp-Asp-Arg-Val-His-Pro-Val-His- Ala-X (III) (92) Y-Asp-Arg-Val-His-Pro-Val-His-Ala-Gly-Pro- Ile-Ala-Pro-Gly-Gln-X (IV) (90) Y-Tyr-Ser-Pro-Thr-Ser-Ile-Leu-Asp-Ile-Arg- Gln-Gly-Pro-Lys-Glu-Pro-Phe-Arg-Asp-Tyr-Val- Asp-Arg-Phe-Tyr-Lys-Thr-Leu-Arg-z-x (V) (88) Y-Asn-Trp-Nor-Thr-Glu-Thr-Leu-Leu-Val-Gln- Asn-Ala-Asn-Pro-Asp-Cys-Lys-Thr-Ile-Leu-Lys- Ala-Leu-Gly-Pro-Ala-Ala-Thr-Leu-Glu-Glu-Nor- Nor-Thr-Ala-Cys-X (VI) (97) Y-Arg-Glu-Leu-Glu-Arg-Phe-Ala-Val-Asn-Pro-Gly- Leu-Leu-Glu-Thr-Ser-Glu-Gly-Cys-Arg-Gln-Ile- Leu-Gly-Gln-Leu-Gln-Pro-Ser-Leu-Gln-Thr-X (VII) (71) Y-Asp-Thr-Gly-His-Ser-Ser-Gln-Val-Ser-Gln- Asn-Tyr (VIII) (36) Val-Lys-Ile-Glu-Pro-Leu-Gly-Val-Ala-Pro- Thr-Lys-Ala-Lys-Arg-Arg-Val-Val-Gln-Arg Gln-Lys-Arg-Ala-Z-X (IX) (56) Ile-Lys-Gln-Leu-Gln-Ala-Arg-Ile-Leu- Ala-Val-Glu-Arg-Tyr-Leu-Lys-Asp-Gln-Gln-Z-X (X) (39) Arg-Ile-Leu-Ala-Val-Glu-Arg-Tyr-Leu-Lys- Asp-Gln-Gln-Leu-Leu-Gly-Ile-Trp-Gly-Cys- Ser-Gly-Lys-Leu-Ile-Cys-X (XI) (40) Y-Lys-Ser-Leu-Glu-Gln-Ile-Trp-Asn-Asn- Met-Thr-Trp-Met-Glu-Trp-Asp-Arg-Glu- Ile-Asn-Z-X (XII) (23) Y-His-Ser-Leu-Ile-Glu-Glu-Ser-Gln-Asn- Gln-Gln-Glu-Lys-Asn-Glu-Gln-Glu-Leu-Leu- Glu-Leu-Asp-Lys-Trp-Z-x (XIII) (79) Y-Lys-Asp-Gln-Gln-Leu-Leu-Gly-Ile-Trp-Gly- Cys-Ser-Gly-Lys-Leu-Ile-Cys-X,
where X is OH or NH2, and Y and Z, if present, are amino acids added to facilitate coupling, and said peptides are conjugated to a protein for which antibodies in human sera are substantially absent or unconjugated to a protein;
incubating for a sufficient time for complex formation to occur; and
determining the formation of complex as a result of change in a detectable signal resulting from complex formation.
16. A method according to claim 15, wherein said label is a fluorescer.
17. A method according to claim 16, wherein complex formation is determined by fluorescence polarization.
18. A method according to claim 15, wherein said label is an enzyme.
19. A method according to claim 18, wherein complex formation is determined as a result of enzyme activity modulation.
20. A peptide of the formula:
(I) (15) Y-Asp-Cys-Lys-Thr-Ile-Leu-Lys-Ala-Leu- Gly-Pro-Ala-Ala-Thr-Leu-Glu-Glu-Met-Met- Thr-Ala-Cys-X,
where X is OH or NH2, and Y, if present, is an amino acid added to facilitate coupling, N-terminal acetylated I, and I linked to a peptide or protein of at least 5,000 molecular weight, which peptide or protein does not normally bind to antibodies present in a human host.
21. A peptide of the formula:
(II) (17) Y-Leu-Lys-Glu-Thr-Ile-Asn-Glu-Glu-Ala Ala-Glu-Trp-Asp-Arg-Val-His-Pro-Val-His Ala-X,
where X is OH or NH2, and Y, if present, is an amino acid added to facilitate coupling, N-terminal acetylated II, and II linked to a peptide or protein of at least 5,000 molecular weight, which peptide or protein does not normally bind to antibodies present in a human host.
22. A peptide of the formula:
(III) (92) Y-Asp-Arg-Val-His-Pro-Val-His-Ala-Gly-Pro- Ile-Ala-Pro-Gly-Gln-X,
where X is OH or NH2, and Y, if present, is an amino acid added to facilitate coupling, N-terminal acetylated III, and III linked to a peptide or protein of at least 5,000 molecular weight, which peptide or protein does not normally bind to antibodies present in a human host.
23. A peptide of the formula:
(IV) (90) Y-Tyr-Ser-Pro-Thr-Ser-Ile-Leu-Asp-Ile-Arg Gln-Gly-Pro-Lys-Glu-Pro-Phe-Arg-Asp-Tyr-Val- Asp-Arg-Phe-Tyr-Lys-Thr-Leu-Arg-Z-X,
where X is OH or NH2, and Y, if present, is an amino acid added to facilitate coupling, N-terminal acetylated IV, and IV linked to a peptide or protein of at least 5,000 molecular weight, which peptide or protein does not normally bind to antibodies present in a human host.
24. A peptide of the formula:
(V) (88) Y-Asn-Trp-Nor-Thr-Glu-Thr-Leu-Leu-Val-Gln- Asn-Ala-Asn-Pro-Asp-Cys-Lys-Thr-Ile-Leu-Lys- Ala-Leu-Gly-Pro-Ala-Ala-Thr-Leu-Glu-Glu-Nor- Nor-Thr-Ala-CyS-X,
where X is OH or NH2, and Y, if present, is an amino acid added to facilitate coupling, N-terminal acetylated V, and V linked to a peptide or protein of at least 5,000 molecular weight, which peptide or protein does not normally bind to antibodies present in a human host.
25. A peptide of the formula:
(VI) (97) Y-Arq-Glu-Leu-Glu-Arg-Phe-Ala-Val-Asn-Pro-Gly- Leu-Leu-Glu-Thr-Ser-Glu-Gly-Cys-Arg-Gln-Ile- Leu-Gly-Gln-Leu-Gln-Pro-Ser-Leu-Gln-Thr-X,
where X is OH or NH2, and Y, if present, is an amino acid added to facilitate coupling, N-terminal acetylated VI, and VI linked to a peptide or protein of at least 5,000 molecular weight, which peptide or protein does not normally bind to antibodies present in a human host.
26. A peptide of the formula:
(VII) (71) Y-Asp-Thr-Gly-His-Ser-Ser-Gln-Val-Ser-Gln- Asn-Tyr
where Y, if present, is an amino acid added to facilitate coupling, N-terminal acetylated VII, and VII linked to a peptide or protein of at least 5,000 molecular weight, which peptide or protein does not normally bind to antibodies present in a human host.
27. A peptide of the formula:
(VIII) (36) Val-Lys-Ile-Glu-Pro-Leu-Gly-Val-Ala-Pro- Thr-Lys-Ala-Lys-Arg-Arg-Val-Val-Gln-Arg- Glu-Lys-Arg-Ala-Z-X,
where X is OH or Nil2, and Z, if present, is an amino acid added to facilitate coupling, N-terminal acetylated VIII, and VIII linked to a peptide or protein of at least 5,000 molecular weight, which peptide or protein does not normally bind to antibodies present in a human host.
28. A peptide of the formula:
(IX) (56) Ile-Lys-Gln-Leu-Gln-Ala-Arg-Ile-Leu- Ala-Val-Glu-Arg-Tyr-Leu-Lys-Asp-Gln-Gln-Z-X,
where X is OH or NH2, and Z, if present, is an amino acid added to facilitate coupling, N-terminal acetylated IX, and IX linked to a peptide or protein of at least 5,000 molecular weight, which peptide or protein does not normally bind to antibodies present in a human host.
29. A peptide of the formula:
(X) (39) Arg-Ile-Leu-Ala-Val-Glu-Arg-Tyr-Leu-Lys- Asp-Gln-Gln-Leu-Leu-Gly-Ile-Trp-Gly-Cys- Ser-Gly-Lys-Leu-Ile-Cys-X,
where X is OH or NH2, N-terminal acetylated X, and X linked to a peptide or protein of at least 5,000 molecular weight, which peptide or protein does not normally bind to antibodies present in a human host.
30. A peptide of the formula:
(XI) (40) Y-Lys-Ser-Leu-Glu-Gln-Ile-Trp-Asn-Asn- Met-Thr-Trp-Met-Glu-Trp-Asp-Arg-Glu- Ile-Asn-Z-X,
where X is OH or NH2, and each of Y and Z, if present, is an amino acid added to facilitate coupling, N-terminal acetylated XI, and XI linked to a peptide or protein of at least 5,000 molecular weight, which peptide or protein does not normally bind to antibodies present in a human host.
31. A peptide of the formula:
(XII) (23) Y-His-Ser-Leu-Ile-Glu-Glu-Ser-Gln-Asn- Gln-Gln-Glu-Lys-Asn-Glu-Gln-Glu-Leu-Leu- Glu-Leu-Asp-Lys-Trp-Z-X,
where X is OH or NH2, and each of Y and Z, if present, is an amino acid added to facilitate coupling, N-terminal acetylated XII, and XII linked to a peptide or protein of at least 5,000 molecular weight, which peptide or protein does not normally bind to antibodies present in a human host.
32. A peptide of the formula:
(XIII) (79) Y-Lys-Asp-Gln-Gln-Leu-Leu-Gly-Ile-Trp-Gly- Cys-Ser-Gly-Lys-Leu-Ile-Cys-X,
where X is OH or NH2, and each of Y and Z. If present, is an amino acid added to facilitate coupling, N-terminal acetylated XIII, and XIII linked to a peptide or protein of at least 5,000 molecular weight, which peptide or protein does not normally bind to antibodies present in a human host.
33. A vaccine composition comprising at least one peptide of the peptides:
(I) (15) Y-Asp-Cys-Lys-Thr-Ile-Leu-Lys-Ala-Leu- Gly-Pro-Ala-Ala-Thr-Leu-Glu-Glu-Met-Met Thr-Ala-Cys-X (II) (17) Y-Leu-Lys-Glu-Thr-Ile-Asn-Glu-Glu-Ala- Ala-Glu-Trp-Asp-Arg-Val-His-Pro-Val-His- Ala-Z-X (III) (92) Y-Asp-Arg-Val-His-Pro-Val-His-Ala-Gly-Pro- Ile-Ala-Pro-Gly-Gln-X (IV) (90) Y-Tyr-Ser-Pro-Thr-Ser-Ile-Leu-Asp-Ile-Arg- Gln-Gly-Pro-Lys-Glu-Pro-Phe-Arg-Asp-Tyr-Val Asp-Arg-Phe-Tyr-Lys-Thr-Leu-Arg-Z-x (V) (88) Y-Asn-Trp-Nor-Thr-Glu-Thr-Leu-Leu-Val-Gln- Asn-Ala-Asn-Pro-Asp-Cys-Lys-Thr-Ile-Leu-Lys- Ala-Leu-Gly-Pro-Ala-Ala-Thr-Leu-Glu-Glu-Nor- Nor-Thr-Ala-Cys-X (VI) (97) Arg-Glu-Leu-Glu-Arg-Phe-Ala-Val-Asn-Pro-Gly- Leu-Leu-Glu-Thr-Ser-Glu-Gly-Cys-Arg-Gln-Ile- Leu-Gly-Gln-Leu-Gln-Pro-Ser-Leu-Gln-Thr (VII) (71) Asp-Thr-Gly-His-Ser-Ser-Gln-Val-Ser-Gln-Asn-Tyr (VIII) (36) Val-Lys-Ile-Glu-Pro-Leu-Gly-Val-Ala-Pro- Thr-Lys-Ala-Lys-Arg-Arg-Val-Val-Gln-Arg- Glu-Lys-Arg-Ala-Z-X (IX) (56) Ile-Lys-Gln-Leu-Gln-Ala-Arg-Ile-Leu- Ala-Val-Glu-Arg-Tyr-Leu-Lys-Asp-Gln-Gln-Z-X (X) (39) Arg-Ile-Leu-Ala-Val-Glu-Arg-Tyr-Leu-Lys- Asp-Gln-Gln-Leu-Leu-Gly-Ile-Trp-Gly-Cys- Ser-Gly-Lys-Leu-Ile-Cys-X (XI) (40) Y-Lys-Ser-Leu-Glu-Gln-Ile-Trp-Asn-Asn- Met-Thr-Trp-Met-Glu-Trp-Asp-Arg-Glu- Ile-Asn-Z-X (XII) (23) Y-His-Ser-Leu-Ile-Glu-Glu-Ser-Gln-Asn- Gln-Gln-Glu-Lys-Asn-Glu-Gln-Glu-Leu-Leu- Glu-Leu-Asp-Lys-Trp-Z-X (XIII) (79) Lys-Asp-Gln-Gln-Leu-Leu-Gly-Ile-Trp-Gly- Cys-Ser-Gly-Lys-Leu-Ile-Cys,
where X is OH or NH2, and each of Y and Z. when present, are amino acids added to facilitate coupling,
or said peptides conjugated to an immunogenic protein, said peptides or conjugates being present in an amount to provide an immunogenic response in a physiologically acceptable carrier.
34. A vaccine according to claim 34, wherein said peptides are coupled to immunogenic proteins.
35. A vaccine according to claim 34, wherein at least one peptide is I to VII and at least one peptide is VIII to XIII.
36. A vaccine according to any of claims 33, 34 or 35, wherein said immunogenic protein is tetanus toxoid.
US10305271 1985-04-29 2002-11-26 Synthetic antigen for the detection of aids-related disease Abandoned US20030211117A1 (en)

Priority Applications (4)

Application Number Priority Date Filing Date Title
US72805285 true 1985-04-29 1985-04-29
US06767303 US4629783A (en) 1985-04-29 1985-08-19 Synthetic antigen for the detection of AIDS-related disease
US07541163 US6485900B1 (en) 1985-04-29 1990-06-20 Synthetic antigen for the detection of AIDS-related disease
US10305271 US20030211117A1 (en) 1985-04-29 2002-11-26 Synthetic antigen for the detection of aids-related disease

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
US10305271 US20030211117A1 (en) 1985-04-29 2002-11-26 Synthetic antigen for the detection of aids-related disease

Related Parent Applications (1)

Application Number Title Priority Date Filing Date
US07541163 Continuation US6485900B1 (en) 1985-04-29 1990-06-20 Synthetic antigen for the detection of AIDS-related disease

Publications (1)

Publication Number Publication Date
US20030211117A1 true true US20030211117A1 (en) 2003-11-13

Family

ID=27111625

Family Applications (2)

Application Number Title Priority Date Filing Date
US06767303 Expired - Lifetime US4629783A (en) 1985-04-29 1985-08-19 Synthetic antigen for the detection of AIDS-related disease
US10305271 Abandoned US20030211117A1 (en) 1985-04-29 2002-11-26 Synthetic antigen for the detection of aids-related disease

Family Applications Before (1)

Application Number Title Priority Date Filing Date
US06767303 Expired - Lifetime US4629783A (en) 1985-04-29 1985-08-19 Synthetic antigen for the detection of AIDS-related disease

Country Status (3)

Country Link
US (2) US4629783A (en)
CA (1) CA1341605C (en)
ES (1) ES8708058A1 (en)

Families Citing this family (166)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4525300A (en) * 1983-01-07 1985-06-25 Japanese Foundation For Cancer Research Human leukemia virus-related peptides, antibodies of the peptides and a process for production of the antibodies
US7045130B1 (en) 1984-10-18 2006-05-16 Institut Pasteur Antibodies against antigens of human immunodeficiency virus (HIV-1)
US5374519A (en) * 1983-12-05 1994-12-20 Institut Pasteur Oligopeptides comprising p18 protein of human immunodeficiency virus (HIV), compositions comprising peptides of p18 protein of HIV, and diagnostic kits and methods for detecting acquired immune deficiency syndrome (AIDS)
US7232654B1 (en) 1983-09-15 2007-06-19 Institut Pasteur DNA sequence of the LTR region of human immunodeficiency virus type 1 (HIV-1)
US6894152B1 (en) 1984-11-16 2005-05-17 Institut Pasteur Cloned DNA sequences related to the genomic RNA of lymphadenopathy-associated-virus (LAV) and proteins encoded by said LAV genomic RNA
US7205102B1 (en) 1983-09-15 2007-04-17 Institut Pasteur Cloned DNA sequences related to the genomic RNA of lymphadenopathy-associated virus (LAV) and proteins encoded by said LAV genomic RNA
US7351412B1 (en) 1983-12-05 2008-04-01 Institut Pasteur Cloned DNA sequences related to the genomic RNA of lymphadenopathy-associated-virus (LAV) and proteins encoded by said LAV genomic RNA
US5217861A (en) * 1983-09-15 1993-06-08 Institut Pasteur Antigen of a human retrovirus, namely p18 protein of human immunodeficiency virus (HIV), compositions containing the antigen, a diagnostic method for detecting acquired immune deficiency syndrome (AIDS) and pre-AIDS and a kit therefor
US5705612A (en) * 1984-10-18 1998-01-06 Institut Pasteur And Centre National De La Recherche Scientifique NEF peptide encoded by human immunodefiency virus type 1 (HIV-1)
US5173400A (en) * 1983-09-15 1992-12-22 Institut Pasteur Antibody detection of antibodies to viral proteins in serum
US5610035A (en) * 1983-12-05 1997-03-11 Institut Pasteur Centre National De La Recherche Scientific Methods for the preparation of hybridomas producing lymphadenopathy-associated virus (LAV) GP110-specific monoclonal antibodies and methods for the purification of GP110 employing said monoclonal antibodies
GB8423659D0 (en) 1984-09-19 1984-10-24 Pasteur Institut Cloned dna sequences
CA1341423C (en) * 1984-10-31 2003-03-04 Paul A. Luciw Recombinant proteins of viruses associated with lymphadenopathy syndrome and/or acquired immune deficiency syndrome
US7273695B1 (en) * 1984-10-31 2007-09-25 Novartis Vaccines And Diagnostics, Inc. HIV immunoassays using synthetic envelope polypeptides
US7408053B1 (en) * 1984-10-31 2008-08-05 Novartis Vaccines And Diagnostics, Inc. Recombinant DNA constructs for GAG polypeptides and polypeptides produced thereby
US7205101B1 (en) * 1984-10-31 2007-04-17 Novartis Vaccines And Diagnostics, Inc. Human immunodeficiency virus (HIV) nucleotide sequences, recombinant polypeptides, and applications thereof
US7285271B1 (en) * 1984-10-31 2007-10-23 Novartis Vaccines And Diagnostics, Inc. Antigenic composition comprising an HIV gag or env polypeptide
GB8429099D0 (en) 1984-11-16 1984-12-27 Pasteur Institut Closed dna sequences
US4774175A (en) * 1985-03-01 1988-09-27 Centocor, Inc. Immunochemical methods for the detection of antibody against HTLV-III
US4784941A (en) * 1985-04-08 1988-11-15 Genetic Systems Corporation Expression and diagnostic use of pENV-3 encoded peptides which are immunologically reactive with antibodies to LAV
FR2580177B2 (en) * 1985-04-15 1989-06-02 Pasteur Institut exposed antigens to glycoprotein of the AIDS virus envelope glycoprotein including precursors of this, processes for obtaining these antigens and means used in these processes, applications of these antigens in the preparation of immunogenic compositions or for the diagnosis of AIDS or conditions are related to him
DK171119B1 (en) * 1985-04-19 1996-06-17 Hoffmann La Roche AIDS virus envelope protein, expression vector carrying the virus coat protein, transformants transformed with the expression vector, method of producing viral coat protein, method for the detection of AIDS antibodies, Method for the determination of AIDS virus, vaccine against AIDS, antibodies to viral coat protein and employing it for preparing a the vaccine or the testnin
US5773210A (en) * 1985-04-19 1998-06-30 The United States Of America As Represented By The Department Of Health And Human Services Acquired immune deficiency syndrome (AIDS) viral envelope protein and method of testing for AIDS
US4629783A (en) * 1985-04-29 1986-12-16 Genetic Systems Corporation Synthetic antigen for the detection of AIDS-related disease
US6130314A (en) * 1986-03-26 2000-10-10 Genetic Systems Corporation Synthetic antigen for the detection of AIDS-related disease
DE3650175D1 (en) * 1985-04-29 1995-02-02 Genetic Systems Corp Synthetic antigens for evidence of AIDS.
US4843011A (en) * 1985-08-01 1989-06-27 Akzo N.V. Monoclonal antibodies for binding HTLV-III proteins, and cell lines for their production
JP2564268B2 (en) * 1985-08-28 1996-12-18 協和醗酵工業株式会社 Fusion antigen polypeptide
US4777127A (en) * 1985-09-30 1988-10-11 Labsystems Oy Human retrovirus-related products and methods of diagnosing and treating conditions associated with said retrovirus
US4956273A (en) * 1985-10-24 1990-09-11 Southwest Foundation For Biomedical Research Synthetic peptides and method of use for diagnosis and vaccination for AIDS and ARC
EP0225066A3 (en) * 1985-11-16 1988-07-13 SPOFA Spojené Podniky Pro Zdravotnickou Vyrobu Antigen determinant peptides and a process for the preparation thereof
US6086880A (en) * 1985-12-26 2000-07-11 The University Of Saskatchewan Rotavirus peptide compositions and methods of use
US5066782A (en) * 1986-01-22 1991-11-19 Institut Pasteur Retrovirus capable of causing AIDS, means and method for detecting it in vitro
US6514691B1 (en) 1986-01-22 2003-02-04 Institut Pasteur Peptides of human immunodeficiency virus type 2 (HIV-2), antibodies against peptides of HIV-2, and methods and kits for detecting HIV-2
US5306614A (en) * 1986-01-22 1994-04-26 Institut Pasteur Methods and kits for diagnosing human immunodeficiency virus type 2(HIV-2)
US4839288A (en) * 1986-01-22 1989-06-13 Institut Pasteur Retrovirus capable of causing AIDS, antigens obtained from this retrovirus and corresponding antibodies and their application for diagnostic purposes
US5310651A (en) * 1986-01-22 1994-05-10 Institut Pasteur DNA probes of human immunodeficiency virus type 2 (HIV-2), and methods employing these probes for dectecting the presence of HIV-2
JP2948823B2 (en) * 1987-01-16 1999-09-13 アンステイテユ・パストウール Human immunodeficiency retrovirus (hiv) associated antigenic peptides, detection and use of the vaccine of the virus-infected
US6037165A (en) 1986-01-22 2000-03-14 Institut Pasteur Methods for the preparation of human immunodeficiency virus type 2 (HIV-2) and antigens encoped thereby
US5976785A (en) * 1986-01-22 1999-11-02 Institut Pasteur Competitive assays for determining the effectiveness of a human immunodeficiency virus type 2 (HIV-2) antiviral agent, employing peptides and proteins of HIV-2
US6054565A (en) 1986-03-03 2000-04-25 Institut Pasteur Nucleic Acids of HIV-2, Diagnostic Test Kit and Method using Nucleic Acid Probes of HIV-2
ES2013295B3 (en) 1986-01-22 1990-05-01 Pasteur Institut Type retroviruses HIV-2 capable of causing AIDS, antigenic and nucleic constituents.
US5580739A (en) * 1986-01-22 1996-12-03 Institut Pasteur Peptides of human immunodeficiency virus type 2 (HIV-2) and in vitro diagnostic methods and kits employing the peptides for the detection of HIV-2
US5364933A (en) 1986-03-03 1994-11-15 Institut Pasteur Methods of immunopurification of antigens of human immunodeficiency virus type 2 (HIV-2)
US6322964B1 (en) * 1987-01-16 2001-11-27 Genetic Systems Corporation Synthetic HIV-2 gag and env oligopeptides reactive with HIV-2 specific antibodies
US5079342A (en) * 1986-01-22 1992-01-07 Institut Pasteur Cloned DNA sequences related to the entire genomic RNA of human immunodeficiency virus II (HIV-2), polypeptides encoded by these DNA sequences and use of these DNA clones and polypeptides in diagnostic kits
US5830641A (en) * 1986-01-22 1998-11-03 Institut Pasteur In vitro diagnostic assays for the detection of HIV-1 or HIV-2 employing viral-specific antigens and antibodies
DE3783237D1 (en) * 1986-01-31 1993-02-11 Ist Naz Stud Cura Dei Tumori Kit and method for determination of the wucherungsaktivitaet of human tumors.
US4772547A (en) * 1986-02-03 1988-09-20 Hoffmann-La Roche Inc. HTLV-III envelope peptides
US4734362A (en) * 1986-02-03 1988-03-29 Cambridge Bioscience Corporation Process for purifying recombinant proteins, and products thereof
US4753873A (en) * 1986-02-03 1988-06-28 Cambridge Bioscience Corporation Peptides for the diagnosis of HTLV-III antibodies, their preparation and use
US4808536A (en) * 1986-02-27 1989-02-28 Centocor, Inc. Immunochemical method for detection of antibody against HTLV-III core protein based upon recombinant HTLV-III gag gene encoded protein
US4879212A (en) * 1986-04-02 1989-11-07 United Biomedical Inc. Peptide composition and method for the detection of antibodies to HTLV-III
DE3788397D1 (en) * 1986-03-24 1994-01-20 Ortho Pharma Corp Synthetic HTLV-III peptide compositions and their use.
US5075211A (en) * 1986-03-26 1991-12-24 Genetic Systems Corporation Synthetic antigen for the detection of AIDS-related disease
US4904581A (en) * 1986-05-06 1990-02-27 Epitope, Inc. Method of detecting AIDS virus infection
US4917998A (en) * 1986-05-06 1990-04-17 Epitope, Inc. Method of detecting AIDS virus infection
DE3789633D1 (en) * 1986-05-27 1994-05-26 Hoffmann La Roche HTLV-III (LAV) ceiling peptides.
US4957737A (en) * 1986-05-27 1990-09-18 Hoffmann-La Roche Inc. HTLV-III (LAV) envelope peptides
US5276016A (en) * 1986-06-03 1994-01-04 The United States Of America As Represented By The Department Of Health And Human Services Small peptides which inhibit binding to T-4 receptors and act as immunogens
US5529983A (en) * 1986-06-03 1996-06-25 The United States Of America As Represented By The Department Of Health And Human Services Use of small peptides in the treatment of psoriasis
US4816387A (en) * 1986-06-06 1989-03-28 Bio-Research Laboratories, Inc. Method for detection of antibodies to HTLV-III and diagnostic test kit useful therewith
US5017688A (en) * 1986-06-12 1991-05-21 Biogen, Inc. Peptides involved in the pathogenesis of HIV infection
US4943627A (en) * 1986-06-12 1990-07-24 Biogen, Inc. Peptides involved in the pathogenesis of HIV infection
JPH01501547A (en) * 1986-06-12 1989-06-01
WO1987007957A1 (en) * 1986-06-20 1987-12-30 The Regents Of The University Of California Low-level detection of human immunodeficiency virus
US5087557A (en) * 1986-06-23 1992-02-11 Genetic Systems Corporation Human monoclonal antibody to lymphadenopathy-associated virus
US5824482A (en) * 1986-06-23 1998-10-20 Institut Pasteur Purification, cloning, and characterization of a novel human immunodeficiency virus LAVMAL
US5030714A (en) 1986-06-23 1991-07-09 Institut Pasteur Variant of LAV viruses
EP0276279A4 (en) * 1986-07-21 1990-01-08 Southwest Found Biomed Res Composition of matter and method of immunizing against viral causative agents of aids and arc.
US5142025A (en) * 1986-08-01 1992-08-25 Repligen Corporation Recombinant HTLV-III proteins and uses thereof
JP3134874B2 (en) 1986-08-21 2001-02-13 ザ・トラステイーズ・オブ・コロンビア・ユニヴアーシテイ・イン・ザ・シテイ・オブ・ニユー・ヨーク Use of the aids treatment of dna and t4 fragment encoding the T cell surface protein t4
US6673896B1 (en) * 1986-08-21 2004-01-06 The Trustees Of Columbia University In The City Of New York Derivatives of soluble T-4
US5126433A (en) * 1986-08-21 1992-06-30 The Trustees Of Columbia University In The City Of New York Soluble forms of the t cell surface protein cd4
US5958678A (en) * 1986-08-21 1999-09-28 The Trustees Of Columbia University In The City Of New York DNA encoding the T cell surface protein T4 and use of fragments of T4 in the treatment of AIDS
US5110906A (en) * 1986-08-21 1992-05-05 The Trustees Of Columbia University In The City Of New York Derivatives of soluble T-4
DE3638767A1 (en) * 1986-11-13 1988-05-26 Behringwerke Ag Lactoferrin-containing process incubation for festphasenimmunometrisches and its use
DE3787002D1 (en) * 1986-12-30 1993-09-16 Us Health Synthetic peptides, cellular immunity against the AIDS virus and proteins produce a result.
US5081226A (en) * 1986-12-30 1992-01-14 The United States Of America As Represented By The Department Of Health And Human Services Synthetic peptides sharing sequence homology with the HIV envelope protein
EP0300031A4 (en) * 1987-01-28 1990-05-14 Ortho Pharma Corp Immunosuppressive peptides and methods of use.
US4861707A (en) * 1987-02-02 1989-08-29 E. I. Du Pont De Nemours And Company Human immunodeficiency virus antigen
US5141867A (en) * 1987-02-02 1992-08-25 E. I. Du Pont De Nemours And Company Nucleotide sequence encoding a human immunodeficiency virus antigen
FR2610632B1 (en) * 1987-02-11 1990-12-21 Pasteur Institut Peptides characteristics of human immunodeficiency retroviruses (hiv virus) applications in the diagnosis of infections with viruses and some of these, if any, to vaccination against AIDS
CA1341436C (en) * 1987-03-25 2003-08-05 Wesley L. Cosand Synthetic antigens for the detection of aids-related disease caused by lav-2
FI881418A (en) * 1987-03-27 1988-09-28 Virovahl Sa Include synthetic Peptide antigener Foer faststaellande of HIV-1 infection.
US5670309A (en) * 1987-04-16 1997-09-23 Johnson & Johnson Methods and diagnostic kits for the detections of HIV-2-specific antibodies employing polypeptides obtained from the simian immunodeficiency virus
EP0309566B1 (en) * 1987-04-16 1994-01-12 Johnson And Johnson Stlv-iii-related polypeptides, diagnostic systems and assay methods
EP0311675A4 (en) * 1987-04-24 1989-10-04 Biogen Nv Immunotherapeutic methods and compositions.
US4921787A (en) * 1987-05-01 1990-05-01 Cambridge Bioscience Corporation Detection of antibodies to human immunodeficiency virus by agglutination of antigen coated latex
US5104790A (en) * 1987-06-29 1992-04-14 Genetic Systems Corporation Monoclonal antibodies to specific antigenic regions of the human immunodeficiency virus and methods for use
EP0324834B1 (en) * 1987-07-13 1995-11-08 Verigen, Inc. Method for rapid and sensitive detection of hiv-i antibodies
DK362287D0 (en) * 1987-07-13 1987-07-13 Kurt Baekgaard Osther Method for rapid and sensitive detection of HIV-1 antibodies
DE3854553T2 (en) * 1987-07-13 1996-03-21 Verigen Inc A method for rapid and sensitive detection of IgM antibodies against retroviral antigens.
US5447837A (en) * 1987-08-05 1995-09-05 Calypte, Inc. Multi-immunoassay diagnostic system for antigens or antibodies or both
JP2902658B2 (en) * 1987-08-21 1999-06-07 スクリップス クリニック アンド リサーチ ファウンデーション Hiv-1-related polypeptides, diagnostic systems such and assay methods
US5086002A (en) * 1987-09-07 1992-02-04 Agen Biomedical, Ltd. Erythrocyte agglutination assay
EP0396559B1 (en) * 1987-10-28 1996-08-14 Ferring AB Hiv peptides, artificial hiv antigens and immunoassay kits
EP0316495A1 (en) * 1987-11-19 1989-05-24 Iaf Biochem International Inc. Synthetic peptides useful for the detection of aids-related antibodies in human sera
WO1989005821A1 (en) * 1987-12-21 1989-06-29 Arch Development Corporation Hiv-related antigens and antibodies
US6214537B1 (en) * 1988-01-27 2001-04-10 Biochem Immunosystems, Inc. Synthetic peptides and mixtures thereof for detecting HIV antibodies
US6210874B1 (en) 1988-01-27 2001-04-03 Biochem Immunosystems, Inc. Synthetic peptides and mixtures thereof for detecting HIV antibodies
US6218102B1 (en) * 1988-01-27 2001-04-17 Biochem Immunosystems, Inc. Synthetic peptides and mixtures thereof for detecting HIV antibodies
US5241047A (en) * 1988-12-08 1993-08-31 Biochem Pharma Inc. Synthetic peptides and mixtures thereof for detecting HIV antibodies
US5763160A (en) * 1988-02-12 1998-06-09 United Biomedical, Inc. Synthetic peptides and process of using same for the detection of antibodies to human immunodeficiency virus (HIV) gp120 envelope protein, diagnosis of AIDS and pre-AIDS conditions and as vaccines
EP0330359A3 (en) * 1988-02-25 1991-06-05 Bio-Rad Laboratories, Inc. Composition useful in the diagnosis and treating of hiv-1 infection
US5562905A (en) * 1988-04-26 1996-10-08 E. I. Du Pont De Nemours And Company Human immunodeficiency virus (hiv) env-coded peptide capable of eliciting hiv-inhibiting antibodies in mammals
DE68925660T2 (en) * 1988-06-10 1996-09-26 United Biomedical Inc Peptide fragments of HIV
WO1990000399A1 (en) * 1988-07-08 1990-01-25 Smithkline Beckman Corporation Retroviral protease binding peptides
US5354846A (en) * 1988-11-18 1994-10-11 Michael Kehoe Streptolysin O antigen derivatives, its production and uses
DE68925909T2 (en) * 1988-12-20 1996-11-14 Clarity Technologies Inc Synthetic HIV-like peptides, their compositions and uses
US5196510A (en) * 1988-12-29 1993-03-23 Cytogen Corporation Molecular recognition units
US5439792A (en) * 1989-06-02 1995-08-08 Genetic Systems Corporation Cysteine thiol-protected peptides for use in immunoassays
US6083504A (en) * 1989-08-24 2000-07-04 Bioclonetics Incorporated Human monoclonal antibodies directed against the transmembrane glycoprotein (GP41) of human immunodeficiency virus-1 (HIV-1)
JP2807287B2 (en) * 1989-10-13 1998-10-08 日本赤十字社 Peptides and use thereof
CA2072351A1 (en) * 1990-01-05 1991-07-06 Andrew J. Mcmichael Hiv-1 core protein fragments
DE69129207T2 (en) * 1990-01-16 1998-10-08 Orgenics Ltd Peptides derived from viral HIV-envelope glycoproteins, their use for the detection of infection of these viruses and for vaccination against AIDS
EP0511216A4 (en) * 1990-01-16 1993-05-05 Prutech Res Dev Partnership Ii Combination assay for antibody or antigen
US5639594A (en) * 1990-02-16 1997-06-17 United Biomedical, Inc. Linear and branched peptides effective in diagnosing and detecting non-A, non-B hepatitis
FR2663034B1 (en) * 1990-06-06 1995-04-21 Neosystem Sa Peptides of the antigen Sm-D and their use, especially for the diagnosis of systemic lupus erythematosus.
US5359029A (en) * 1990-07-18 1994-10-25 Biochem Immunosystems Inc. Peptides and analogues and mixtures thereof for detecting antibodies to HTLV-I and HTLV-II viruses
WO1992003579A1 (en) * 1990-08-16 1992-03-05 Diagnostic Biotechnology, Inc. An augmented western blot format and immunoassay for detection of viral antibodies
DE4039925A1 (en) * 1990-12-14 1992-06-17 Behringwerke Ag Selected peptides of the group specific antigen (gag) of human immunodeficiency virus (HIV), their preparation and use
US5753488A (en) * 1991-05-23 1998-05-19 Martin; William John Isolated stealth viruses and related vaccines
US6685939B2 (en) 1991-08-14 2004-02-03 Genentech, Inc. Method of preventing the onset of allergic disorders
US5994514A (en) * 1991-08-14 1999-11-30 Genentech, Inc. Immunoglobulin variants
US6329509B1 (en) * 1991-08-14 2001-12-11 Genentech, Inc. Anti-IgE antibodies
ES2161228T3 (en) * 1992-02-10 2001-12-01 Baxter Int Method for controlling viral contamination in blood samples.
US7326535B2 (en) 1993-09-15 2008-02-05 Ortho Diagnostic Systems Inc. Immunoreactive peptides from Epstein-Barr virus
US7060283B1 (en) 1992-09-15 2006-06-13 Ortho Diagnostic Systems, Inc. Immunoreactive peptides from Epstein-Barr virus
DE59309207D1 (en) * 1992-10-06 1999-01-21 Dade Behring Marburg Gmbh Retrovirus of the HIV group, and its use
WO1994026306A1 (en) 1993-05-12 1994-11-24 Chiron Corporation Conserved motif of hepatitis c virus e2/ns1 region
US20040259113A1 (en) * 1993-07-22 2004-12-23 Mayo Foundation For Medical Education And Research, Hybritech Incorporated Method for detection of metastatic prostate cancer
US6103237A (en) * 1993-07-22 2000-08-15 Hybritech Incorporated Stable variant hK2 polypeptide
US5516639A (en) * 1993-07-22 1996-05-14 Mayo Foundation For Medical Education And Research Antibodies specific for human prostate glandular kallkrein
DE4405810A1 (en) 1994-02-23 1995-08-24 Behringwerke Ag From a retrovirus of the HIV group derived peptides and their use
CA2189774A1 (en) * 1994-05-10 1995-11-16 Donald J. Tindall Recombinant hk2 polypeptide
US6506553B1 (en) 1995-03-30 2003-01-14 Ortho Diagnostics Systems, Inc. Method for diagnosis of Epstein-Barr virus associated disease
US20030036106A1 (en) * 1996-10-28 2003-02-20 Burke Thomas J. Kinase and phosphatase activity measurement using fluorescence polarization
US6479263B1 (en) 1996-11-14 2002-11-12 Baylor College Of Medicine Method for detection of micrometastatic prostate cancer
US7033589B1 (en) * 1997-02-20 2006-04-25 Biogen Idec Ma Inc. γ-1 anti-human CD23 monoclonal antibodies and use thereof as therapeutics
US6893636B2 (en) * 1997-02-20 2005-05-17 Biogen Idec Ma Inc. Gamma-1 and gamma-3 anti-human CD23 monoclonal antibodies and use thereof as therapeutics
US20030118985A1 (en) * 1997-05-14 2003-06-26 Hunt Jeffrey C. Mouse monoclonal antibody (5-21-3) to human immunodeficiency virus gp41 protein
WO1998059073A1 (en) 1997-06-20 1998-12-30 Mayo Foundation For Medical Education And Research Method for detection of breast cancer
DE69837666T2 (en) 1997-08-01 2007-12-27 Genetic Systems Corp., Redmond Synthetic antigens to detect antibodies having immunoreactivity against hiv virus
JP3255293B2 (en) * 1997-09-05 2002-02-12 松下電器産業株式会社 Fluorescence polarization
US6749856B1 (en) 1997-09-11 2004-06-15 The United States Of America, As Represented By The Department Of Health And Human Services Mucosal cytotoxic T lymphocyte responses
WO1999016467A1 (en) * 1997-09-30 1999-04-08 Pharmacia & Upjohn Diagnostics Ab Non-anaphylactic forms of allergens and their use
JP4805511B2 (en) * 1999-12-23 2011-11-02 インターナショナル エイズ バクシーン イニシアティブ Improvements in the immune response against hiv, or improvements to the immune response
KR100768174B1 (en) * 2000-12-06 2007-10-17 삼성에스디아이 주식회사 Electron gun for cathode ray tube
DE60336978D1 (en) * 2003-04-11 2011-06-16 Pasteur Institut Synthetic peptide HIV vaccine: the CBD epitope as an efficient immunogen for the induction of HIV-neutralizing antibodies
US7727752B2 (en) 2003-07-29 2010-06-01 Life Technologies Corporation Kinase and phosphatase assays
US7619059B2 (en) 2003-07-29 2009-11-17 Life Technologies Corporation Bimolecular optical probes
EP2107374A1 (en) * 2003-09-11 2009-10-07 Idexx Laboratories, Inc. Polypeptide for diagnosis of vaccination
US7285272B2 (en) * 2003-12-18 2007-10-23 Idexx Laboratories, Inc. Method and device for detecting feline immunodeficiency virus
RU2006128593A (en) * 2004-01-07 2008-02-20 Тримерис, Инк. (Us) Synthetic peptides derived region hr2 HIV protein gp41 and their use in therapy for inhibiting human immunodeficiency virus penetration
JP2007523170A (en) * 2004-02-19 2007-08-16 アイデックス ラボラトリーズ インコーポレイテッド Method and apparatus for detection of Feline Immunodeficiency virus
WO2006011919A1 (en) * 2004-06-30 2006-02-02 Idexx Laboratories, Inc. Method and device for detecting feline immunodeficiency virus
US7335360B2 (en) * 2004-06-30 2008-02-26 Idexx Laboratories, Inc. Method and device for detecting feline immunodeficiency virus
JP4662988B2 (en) * 2004-06-30 2011-03-30 アイデックス ラボラトリーズ インコーポレイテッド Method and apparatus for detecting a feline immunodeficiency virus
US20080233566A1 (en) * 2004-08-20 2008-09-25 Bayer Healthcare Llc Methods and materials for detecting mutations in quasispecies having length polymorphisms
US7291338B2 (en) * 2005-03-09 2007-11-06 Idexx Laboratories, Inc. Method and device for detecting feline immunodeficiency virus
JP4682251B2 (en) * 2006-02-02 2011-05-11 トリメリス,インコーポレーテッド hiv fusion inhibitor peptide having improved biological properties
EP2139526A4 (en) * 2007-04-03 2010-07-14 Trimeris Inc Novel formulations for delivery of antiviral peptide therapeutics
WO2009003291A1 (en) * 2007-07-03 2009-01-08 Mount Sinai Hospital Therapeutics and diagnostics for group a streptococci
JP2010540528A (en) * 2007-09-25 2010-12-24 トリメリス,インコーポレーテッド New process for the synthesis of the therapeutic antiviral peptide
EP2553452B1 (en) 2010-04-02 2015-03-04 IDEXX Laboratories, Inc. Detection of feline immunodeficiency virus

Citations (20)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US475896A (en) * 1892-05-31 Cable-grip
US4520113A (en) * 1984-04-23 1985-05-28 The United States Of America As Represented By The Secretary Of The Department Of Health And Human Services Serological detection of antibodies to HTLV-III in sera of patients with AIDS and pre-AIDS conditions
US4629783A (en) * 1985-04-29 1986-12-16 Genetic Systems Corporation Synthetic antigen for the detection of AIDS-related disease
US4708818A (en) * 1983-09-15 1987-11-24 Institut Pasteur Human immunodeficiency viruses associated with Acquired Immune Deficiency Syndrome (AIDS), a diagnostic method for AIDS and pre-AIDS, and a kit therefor
US4753873A (en) * 1986-02-03 1988-06-28 Cambridge Bioscience Corporation Peptides for the diagnosis of HTLV-III antibodies, their preparation and use
US4772547A (en) * 1986-02-03 1988-09-20 Hoffmann-La Roche Inc. HTLV-III envelope peptides
US4774175A (en) * 1985-03-01 1988-09-27 Centocor, Inc. Immunochemical methods for the detection of antibody against HTLV-III
US4784941A (en) * 1985-04-08 1988-11-15 Genetic Systems Corporation Expression and diagnostic use of pENV-3 encoded peptides which are immunologically reactive with antibodies to LAV
US4803156A (en) * 1985-09-11 1989-02-07 New York Blood Center Peptide-beta-lactamase conjugates for enzyme-linked immunoassays
US4808536A (en) * 1986-02-27 1989-02-28 Centocor, Inc. Immunochemical method for detection of antibody against HTLV-III core protein based upon recombinant HTLV-III gag gene encoded protein
US4839288A (en) * 1986-01-22 1989-06-13 Institut Pasteur Retrovirus capable of causing AIDS, antigens obtained from this retrovirus and corresponding antibodies and their application for diagnostic purposes
US4843011A (en) * 1985-08-01 1989-06-27 Akzo N.V. Monoclonal antibodies for binding HTLV-III proteins, and cell lines for their production
US4861707A (en) * 1987-02-02 1989-08-29 E. I. Du Pont De Nemours And Company Human immunodeficiency virus antigen
US4879212A (en) * 1986-04-02 1989-11-07 United Biomedical Inc. Peptide composition and method for the detection of antibodies to HTLV-III
US4943628A (en) * 1988-06-13 1990-07-24 Ortho Pharmaceutical Corporation HIV peptide-inducted T cell stimulation
US4956273A (en) * 1985-10-24 1990-09-11 Southwest Foundation For Biomedical Research Synthetic peptides and method of use for diagnosis and vaccination for AIDS and ARC
US5017688A (en) * 1986-06-12 1991-05-21 Biogen, Inc. Peptides involved in the pathogenesis of HIV infection
US5034511A (en) * 1987-04-13 1991-07-23 Institut Pasteur Variant of LAV viruses
US5156949A (en) * 1984-10-31 1992-10-20 Chiron Corporation Immunoassays for antibody to human immunodeficiency virus using recombinant antigens
US6214539B1 (en) * 1985-04-29 2001-04-10 Bio-Rad Laboratories Synthetic antigen the detection of aids-related disease

Patent Citations (20)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US475896A (en) * 1892-05-31 Cable-grip
US4708818A (en) * 1983-09-15 1987-11-24 Institut Pasteur Human immunodeficiency viruses associated with Acquired Immune Deficiency Syndrome (AIDS), a diagnostic method for AIDS and pre-AIDS, and a kit therefor
US4520113A (en) * 1984-04-23 1985-05-28 The United States Of America As Represented By The Secretary Of The Department Of Health And Human Services Serological detection of antibodies to HTLV-III in sera of patients with AIDS and pre-AIDS conditions
US5156949A (en) * 1984-10-31 1992-10-20 Chiron Corporation Immunoassays for antibody to human immunodeficiency virus using recombinant antigens
US4774175A (en) * 1985-03-01 1988-09-27 Centocor, Inc. Immunochemical methods for the detection of antibody against HTLV-III
US4784941A (en) * 1985-04-08 1988-11-15 Genetic Systems Corporation Expression and diagnostic use of pENV-3 encoded peptides which are immunologically reactive with antibodies to LAV
US4629783A (en) * 1985-04-29 1986-12-16 Genetic Systems Corporation Synthetic antigen for the detection of AIDS-related disease
US6214539B1 (en) * 1985-04-29 2001-04-10 Bio-Rad Laboratories Synthetic antigen the detection of aids-related disease
US4843011A (en) * 1985-08-01 1989-06-27 Akzo N.V. Monoclonal antibodies for binding HTLV-III proteins, and cell lines for their production
US4803156A (en) * 1985-09-11 1989-02-07 New York Blood Center Peptide-beta-lactamase conjugates for enzyme-linked immunoassays
US4956273A (en) * 1985-10-24 1990-09-11 Southwest Foundation For Biomedical Research Synthetic peptides and method of use for diagnosis and vaccination for AIDS and ARC
US4839288A (en) * 1986-01-22 1989-06-13 Institut Pasteur Retrovirus capable of causing AIDS, antigens obtained from this retrovirus and corresponding antibodies and their application for diagnostic purposes
US4772547A (en) * 1986-02-03 1988-09-20 Hoffmann-La Roche Inc. HTLV-III envelope peptides
US4753873A (en) * 1986-02-03 1988-06-28 Cambridge Bioscience Corporation Peptides for the diagnosis of HTLV-III antibodies, their preparation and use
US4808536A (en) * 1986-02-27 1989-02-28 Centocor, Inc. Immunochemical method for detection of antibody against HTLV-III core protein based upon recombinant HTLV-III gag gene encoded protein
US4879212A (en) * 1986-04-02 1989-11-07 United Biomedical Inc. Peptide composition and method for the detection of antibodies to HTLV-III
US5017688A (en) * 1986-06-12 1991-05-21 Biogen, Inc. Peptides involved in the pathogenesis of HIV infection
US4861707A (en) * 1987-02-02 1989-08-29 E. I. Du Pont De Nemours And Company Human immunodeficiency virus antigen
US5034511A (en) * 1987-04-13 1991-07-23 Institut Pasteur Variant of LAV viruses
US4943628A (en) * 1988-06-13 1990-07-24 Ortho Pharmaceutical Corporation HIV peptide-inducted T cell stimulation

Also Published As

Publication number Publication date Type
ES554453D0 (en) grant
ES8708058A1 (en) 1987-11-16 application
CA1341605C (en) 2010-05-18 grant
ES554453A0 (en) 1987-09-01 application
US4629783A (en) 1986-12-16 grant

Similar Documents

Publication Publication Date Title
Wang et al. Detection of antibodies to human T-lymphotropic virus type III by using a synthetic peptide of 21 amino acid residues corresponding to a highly antigenic segment of gp41 envelope protein
Pfaff et al. Antibodies against a preselected peptide recognize and neutralize foot and mouth disease virus.
Earl et al. Native oligomeric human immunodeficiency virus type 1 envelope glycoprotein elicits diverse monoclonal antibody reactivities.
Broder et al. Antigenic implications of human immunodeficiency virus type 1 envelope quaternary structure: oligomer-specific and-sensitive monoclonal antibodies
US5019387A (en) Production of antibodies to HIV
US6261569B1 (en) Retro-, inverso- and retro-inverso synthetic peptide analogues
US5304466A (en) HIV-3 retrovirus and its use
US4983387A (en) HIV related peptides, immunogenic antigens, and use therefor as subunit vaccine for AIDS virus
US5013548A (en) Production of antibodies to HIV
EP0328403A2 (en) Synthetic peptides related to the HIV-GP120-env-protein, and their use
Norrby et al. Discrimination between antibodies to HIV and to related retroviruses using site-directed serology
US4879212A (en) Peptide composition and method for the detection of antibodies to HTLV-III
Papsidero et al. Human immunodeficiency virus type 1-neutralizing monoclonal antibodies which react with p17 core protein: characterization and epitope mapping.
US5166050A (en) Monoclonal antibodies and peptides useful in treating and diagnosing HIV infections
US6458528B1 (en) Diagnosis of feline immunodeficiency virus infection using ENV/GAG polypeptide markers
EP0339504A2 (en) Human immunodeficiency virus (HIV) env-coded peptide capable of eliciting HIV-inhibiting antibodies in mammals
US6153377A (en) Synthetic DNA derived recombinant HIV antigens
US5759769A (en) Tandem synthetic HIV-1 peptides
US5260189A (en) Synthetic HIV-like peptides their compositions and uses
US4784941A (en) Expression and diagnostic use of pENV-3 encoded peptides which are immunologically reactive with antibodies to LAV
US5652342A (en) Heterofunctional cellular immunological reagents, vaccines containing same and methods for the use of same
EP0201716A2 (en) Expression of immunologically reactive viral proteins
US4753873A (en) Peptides for the diagnosis of HTLV-III antibodies, their preparation and use
WO1986002383A1 (en) Envelope antigens of lymphadenopathy associated virus and their applications
Lenzi et al. Liver cytosolic 1 antigen-antibody system in type 2 autoimmune hepatitis and hepatitis C virus infection.