WO2009153731A1 - Système et procédé permettant d'évaluer la qualité d'un échantillon - Google Patents

Système et procédé permettant d'évaluer la qualité d'un échantillon Download PDF

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Publication number
WO2009153731A1
WO2009153731A1 PCT/IB2009/052553 IB2009052553W WO2009153731A1 WO 2009153731 A1 WO2009153731 A1 WO 2009153731A1 IB 2009052553 W IB2009052553 W IB 2009052553W WO 2009153731 A1 WO2009153731 A1 WO 2009153731A1
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information
inf
biological sample
diag
analyzing unit
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PCT/IB2009/052553
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English (en)
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Alphons A. M. L. Bruekers
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Koninklijke Philips Electronics N.V.
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Publication of WO2009153731A1 publication Critical patent/WO2009153731A1/fr

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    • G—PHYSICS
    • G16—INFORMATION AND COMMUNICATION TECHNOLOGY [ICT] SPECIALLY ADAPTED FOR SPECIFIC APPLICATION FIELDS
    • G16B—BIOINFORMATICS, i.e. INFORMATION AND COMMUNICATION TECHNOLOGY [ICT] SPECIALLY ADAPTED FOR GENETIC OR PROTEIN-RELATED DATA PROCESSING IN COMPUTATIONAL MOLECULAR BIOLOGY
    • G16B50/00—ICT programming tools or database systems specially adapted for bioinformatics

Definitions

  • the present invention relates to a system for extraction biological information from of a biological sample.
  • the present invention relates to a system for quality control of a biological sample obtained from a subject prior the extraction of further biological information from said biological sample by a subsequent step(s) of analysis.
  • the invention also relates to an analyzing unit, a method, a corresponding computer program product of the method, and a quality control database.
  • RNA is rapidly degraded from the sample if stored improperly. Authentication of the integrity of a biological may be highly relevant for example in drug abuse programs and anti- doping program.
  • DNA is used either for identification or for diagnostics in certified laboratories where privacy and consistency are guaranteed by procedures rather than by technology. Forensic labs are only interested in checking an identity and are not equipped for medical diagnostics. Moreover the law of many countries is very strict in what data may or may not be retrieved from DNA. In some countries it already is, or soon will be, allowed to analyze DNA for information that could also have been obtained when the person was seen at close distance e.g. color of eyes and hair, ethnicity and body length. DNA-based diagnostics in most cases is in a research phase where anonymous samples are processed or in a hospital environment where privacy is provided in a traditional way.
  • Nuclear DNA is found in basically every cell of a living organism (in humans excepted are erythrocytes) and contains the genetic information used in the development and functioning of all known living organisms and is inherited in a rearranged form from both parents in the process of recombination. Extensive knowledge of in particular nuclear DNA, i.e. its functionality and the DNA sequence of a particular person allows for many applications in the field of healthcare. DNA can e.g. be used for early diagnosis and personalized medicine.
  • Mitochondria are cytoplasmic organelles containing a small, maternally- inherited, circular piece of DNA (mitochondrial DNA or mtDNA) distinct from the linear chromosomes of the nucleus. Thousands of mitochondria can exist in a single cell, and as such there are thousands of copies of mtDNA as well. This is far different then the two copies (maternal and paternal) of a DNA locus that are examined in most forensic analyses. Since mtDNA is maternally-inherited no process of recombination is involved at this stage. Thus unlike nuclear DNA, there is usually no change in mtDNA from parent to offspring. Although mtDNA also recombines, it does so with copies of itself within the same mitochondrion.
  • mtDNA is a powerful tool for tracking ancestry through females (matrilineage) and has been used in this role to track the ancestry of many species back hundreds of generations. Both nuclear DNA and mtDNA can be used to identify individuals by forensic analysis.
  • RNA is the product of the process of transcription of the genetic code of a gene into for example an mRNA carrying the code for subsequent translation into a polypeptide.
  • the advances in medical diagnostics include methods of analysis for the determination of the presence or absence of specific RNA transcript known to correlate with specific clinical conditions in a subject.
  • RNA/DNA analysis it may be important to know whether material is obtained directly from the body (e.g. fresh blood or buccal cells) and is intended to be used for analysis, or is extracted from body-dust (e.g. body fluid stains, lost hair, fingernail, cigarette butt).
  • body-dust e.g. body fluid stains, lost hair, fingernail, cigarette butt.
  • RNA/DNA analysis may be controlled by a PIN (personal identification number) code access control system.
  • PIN personal identification number
  • US patent application 2001/0012611 discloses such a system with an anonymous test kit for example for HIV testing. By coupling a personal code with a specific test kit, anonymity is conserved.
  • the anonymous testing system comprises means for taking a sample of body fluid to be tested, the sample is preferably acquired in private and sent for analyzation to obtain results.
  • the system comprises a test kit for creating a sample of body fluid, a personal code for anonymously identifying the sample and the person, and an electronic file telephonically created and accessed by the person taking the test and identified by the personal code.
  • this system does not prevent a third party, who has obtained, in unauthorized manner, a biological sample (e.g. a hair or some blood), from performing a diagnostic test on the biological sample. Rather, the system may protect the privacy of the person who requests the diagnostic result.
  • a biological sample e.g. a hair or some blood
  • an employer or an insurance company may be tempted to perform unauthorized diagnostic testing relevant for the condition and/or medical well being of an individual.
  • an improved system for controlling, and thus if necessary preventing, the analysis processing and extraction of biological information from samples, which is not obtained directly from the origin of the sample e.g. the body is desirable.
  • an object of the present invention relates to a system for processing information from a biological sample.
  • one aspect of the invention relates to a system for processing information obtained from a biological sample of a subject , the system comprising: - an analyzing unit (AU) for analyzing information obtainable from said biological sample, the analyzing unit being adapted for extracting at least a first (QC-INF) and a second (DIAG-INF) information from said biological sample: a) the first information (QC-INF) being related to the quality of said biological sample, the first information being extracted by a method of quality control analysis, and b) the second information (DIAG-INF) being related to the health and/or condition and/or identity verification of said subject,
  • QC-DB quality control database
  • AU analyzing unit
  • Q-P quality control parameters
  • AF authorization flag
  • threshold parameter for prompting said authorization flag (AF)
  • a processing unit arranged for receiving the second information (DIAG-INF) from the analyzing unit (AU) and processing the second information into third information (RES), wherein the processing of the second information (DIAG-INF) into third information (RES) is dependent on the prompting of the authorization flag (AF) of said biological sample, and/or wherein accessing the second information (DIAG-INF) and/or accessing the third information (RES) is dependent on the prompting of the authorization flag (AF) of said biological sample, and/or wherein the execution of the analysis performed by the analyzing unit (AU) for the extraction of the second information (DIAG-INF) is dependent on the prompting of the authorization flag (AF) of said biological sample.
  • PU processing unit
  • the system may be further provide a receiving unit (RU) for receiving and preparing the biological sample, the analyzing unit (AU) being arranged for co-operation with the receiving unit.
  • RU receiving unit
  • AU analyzing unit
  • the first information (QC-INF) related to the quality of said biological sample may comprise
  • - information being extracted by analysis of genetic material comprising DNA or RNA or comprises - information being extracted by analysis of non-genetic biological material comprising the physical state or level of one or more proteins or polypeptides or peptides, or the biological state of cells or tissue, or any combination of the aforementioned.
  • the quality of said biological sample may defined by the integrity of the biological matter from which the information is being extracted.
  • the integrity of genetic material may be defined by the state of degradation of said genetic material.
  • the integrity of proteins or polypeptides or peptides may defined by the state of degradation or posttranslational modification associated with degradation of said proteins or polypeptides or peptides.
  • the integrity of cells or tissue may defined by the state of cyto logical markers for cellular integrity comprising markers for cell membrane integrity, apoptosis and necrosis.
  • Methods of analysis used for the extraction of the first information (QC-INF) from the biological sample comprises nucleotide sequence amplification, a microarray, cytometry, or electrophoresis.
  • the second information (DIAG-INF) related to the health and/or condition and/or identity verification of said subject may comprises
  • - information being extracted by analysis of genetic material comprising DNA or RNA or comprises - information being extracted by analysis of non-genetic biological material comprising the physical state or level of one or more proteins or polypeptides or peptides, or the biological state of cells or tissue, or any combination of the aforementioned.
  • the first information (QC-INF) and the second information (DIAG-INF) may comprise overlapping information.
  • the system may be integrated in one single unit, which may be portable.
  • Application of the system to a single integrated unit such as a portable unit for example a handheld self-operated system may provide an intelligent management of the consumption of reagents, which may extend the operational times-span between services such as refilling the system with reagents.
  • Such system may further provide a checkpoint for the quality of the biological sample applied to the system.
  • the system provides means of controlling that the sample is not in a state of degradation, which may compromise extraction of information from said sample.
  • the system further provides means of avoiding the application of samples collected from subjects without there knowledge and approval, such as body dust (e.g. hair, blood stains).
  • a second aspect of the invention relates to a quality control database (QC-DB) for cooperating in processing information obtained from a biological sample of a subject, the database being adapted to cooperate with an associated analyzing unit
  • the analyzing unit for analyzing information obtainable from said biological sample, the analyzing unit being adapted for extracting at least a first (QC-INF) and a second (DIAG-INF) information from said biological sample: a) the first information (QC-INF) being related to the quality of said biological sample, the first information being extracted by a method of quality control analysis, and b) the second information (DIAG-INF) being related to the health and/or condition and/or identity verification of said subject, the quality control database (QC-DB) being arranged to receive the first information (QC-INF) from the analyzing unit (AU), the quality control database comprising corresponding sets of quality control parameters (QC-P), an authorization flag (AF) for said biological sample, and a threshold parameter for prompting said authorization flag (AF), the database further being adapted to cooperate with an associated processing unit (PU) arranged for receiving the second information (DIAG-INF) from the analyzing unit (AU) and processing the second information into third information (RES), wherein the processing of the second information (DIA
  • a third aspect of the invention relates to an analyzing unit (AU) for cooperating in processing information obtained from a biological sample of a subject, the analyzing unit (AU) being arranged for analyzing information obtainable from said biological sample, the analyzing unit being adapted for extracting at least a first (QC-INF) and a second (DIAG-INF) information from said biological sample: a) the first information (QC-INF) being related to the quality of said biological sample, the first information being extracted by a method of quality control analysis, and b) the second information (DIAG-INF) being related to the health and/or condition and/or identity verification of said subject, the analyzing unit further being adapted to cooperate with an associated quality control database (QC-DB), the database being arranged to receive the first information (QC-INF) from the analyzing unit (AU), the quality control database comprising corresponding sets of quality control parameters (QC-P), an authorization flag (AF) for said biological sample, and a threshold parameter for prompting said authorization flag (AF), the analyzing unit further being
  • a fourth aspect of the present invention relates to a method for processing information obtained from a biological sample of a subject, the method comprising:
  • an analyzing unit for analyzing information obtainable from said biological sample, the analyzing unit being adapted for extracting at least a first (QC- INF) and a second (DIAG-INF) information from said biological sample: a) the first information (QC-INF) being related to the quality of said biological sample, the first information being extracted by a method of quality control analysis, and b) the second information (DIAG-INF) being related to the health and/or condition and/or identity verification of said subject, - providing a quality control database (QC-DB) arranged to receive the first information (QC- INF) from the analyzing unit (AU), the quality control database comprising corresponding sets of quality control parameters (QC-P), an authorization flag (AF) for said biological sample, and a threshold parameter for prompting said authorization flag (AF), and - providing a processing unit (PU) arranged for receiving the second information (DIAG-INF) from the analyzing unit (AU) and processing the second information into third information (RES), wherein the processing of the second
  • a fifth aspect of the invention relates to a computer program product being adapted to enable a computer system comprising at least one computer having data storage means in connection therewith to implement a method according to the fourth aspect of the invention.
  • This aspect of the invention is particularly, but not exclusively, advantageous in that the present invention may be implemented by a computer program product enabling a computer system to perform the operations of the second aspect of the invention.
  • a computer program product may be provided on any kind of computer readable medium, e.g. magnetically or optically based medium, or through a computer based network, e.g. the Internet.
  • the invention is particularly, but not exclusively, advantageous for providing system that may prevent the analysis processing and extraction of biological information from samples, which does not qualify due to degradation of the material. Furthermore, the system may terminate an array of analysis extracting independent sets of information from the biological sample when no further information is required to conclude on the data, for example predicting the risk of specific disease in a subject. Accordingly, the system provides a means of intelligent management of reagent consumption, which may extend the period of which the system operate without expect technical attention. Moreover, the expenses on costly reagents such as antibodies, enzymes, probes such as fluorescent probes, and radioactive isotopes may be significantly reduced.
  • the system may be particularly advantageous if the analytical device performing the extraction of the information is used on location, which are not certified laboratories and/or handled by personnel with no certified training in the of lab equipments. Examples of such system are handheld biosensors for self-operation and system installed in remote areas for self-operation.
  • the first, second, third, fourth and fifth aspect of the present invention may each be combined with any of the other aspects.
  • Fig. 1 shows a system for quality control of a biological sample obtained from a subject prior the extraction of further biological information from said biological sample by a subsequent step(s) of analysis.
  • AU analyzing unit
  • DIDG-INF quality control database
  • Fig. 2 (a) Agarose yield gel result from intact and from degraded DNA samples, and (b) example of an STR DNA-profile of a degraded sample, figure 7.1 p.147 in Forensic DNA Typing: Biology, Technology, Genetics of STR markers, John M. Butler Fig. 3: Two additional examples STR DNA-prof ⁇ les of degraded material. Top is figure 5 and bottom is part of figure 9 from Evaluating Forensic DNA Evidence: Essential Elements of a Competent Defense Review, W.C. Thompson, S. Ford, T. Doom, M. Raymer and D. Krane, Champion Magazine, April 2003 page 16, National Association of criminal Defense Lawyers (NACDL), WWW 1 NACDL 1 OTg.
  • NACDL National Association of criminal Defense Lawyers
  • Fig. 4 For a variety of 23 samples the qPCR degradation ratio and the
  • Fig. 5 Graphical representation of data from the table of Figure 4. For the 23 samples the qPCR degradation ratio is plotted (line with circles). The D 19/Dl 8 ratio is plotted; the average (dashed) and the extremes (bar).
  • Fig. 6 The data for the D 19/Dl 8 ratio from figure 5 is sorted and a threshold is set at 1.
  • Fig. 7 A flow chart of a method according to the present invention.
  • the present invention provide a system and method for extracting sets of information from biological samples in an ordered manner that comprise extracting a first information from the sample, a checkpoint of evaluating the extracted information prior extracting further information from said sample.
  • the extraction of second and/or further information from the samples is dependent on authorization based on the evaluation of the first information.
  • the extraction the second, third and so fourth information is dependent on the evaluation of the previous information in the line of information.
  • a quality control database QC-DB arranged to receive the first information QC-INF from the analyzing unit AU, the quality control database comprising corresponding sets of quality control parameters QC-P, shown with running index from 1 to n in Figure 1, an authorization flag AF for said biological sample 12, and a threshold parameter for prompting said authorization flag AF (more than one kind of flag is possible as shown in Figure 1), and
  • processing unit PU arranged for receiving the second information DIAG- INF from the analyzing unit AU and processing the second information into third information
  • a receiving unit RU is further provided for receiving and preparing the biological sample 12, the analyzing unit AU being arranged for co-operation with the receiving unit.
  • system according to the invention is integrated in one single unit. In a further embodiment, the system is portable.
  • Typical methods for analysis of DNA are methods such as DNA sequencing, SNP genotyping, PCR, Southern blotting, Northern blotting, dot blotting, microarrays, mass spectrometry, restriction analysis, etc.
  • DNA sequencing may be employed to extract detailed information of any desirable genetic bio markers. However, very often it will be more desirable to use methods which allow a higher throughput, or are simpler to use.
  • Such assays could e.g. be PCR, Southern blotting, Northern blotting, dot blotting, microarrays, mass spectrometry, restriction analysis etc.
  • proximity detection systems like FRET (Fluorescence resonance energy transfer) are suitable.
  • the information extracted from the biological sample is genetic information
  • the information can be extracted from mitochondrial DNA, from chromosomal DNA or from both mitochondrial DNA and chromosomal DNA.
  • the genetic information can also be extracted from RNA, e.g. in form of particular sequences (markers) or in the form of levels of one or more RNAs that may reflect a particular condition.
  • Methods for analysis of proteins are e.g. immunoblotting, other antibody based detection systems, proximity bases detection systems like FRET, mass spectrometry, gel electrophoreses (such as SDS-page, both ID and 2D) protein arrays/chips etc.
  • the present invention is not primarily concerned with the development of such methods, but is rather concerned with the control of information that can be extracted using such methods.
  • the first information QC-INF relates to use of genetic markers selected from the group consisting of: VNTR (variable number tandem repeats), STR (short tandem repeats) and SNP (single nucleotide polymorphism).
  • Short tandem repeat (STR) PCR A preferred kind of marker is the short tandem repeats.
  • STR Short Tandem Repeat
  • the STR-markers used with the system of the invention comprise at least one STR marker selected from the group consisting of: CSFlPO, TPOX, D5S818, D7S820, D13S317, D3S1358, VWA, D16S539, D2S1338, A (gender marker Amelogenin), D8S1179, D21S11, D18S51, D19S433, THOl and FGA.
  • STR marker selected from the group consisting of: CSFlPO, TPOX, D5S818, D7S820, D13S317, D3S1358, VWA, D16S539, D2S1338, A (gender marker Amelogenin), D8S1179, D21S11, D18S51, D19S433, THOl and FGA.
  • the number of STR-markers used with the system is selected from the group consisting of least 2 markers, at least 3 markers, at least 4 markers, at least 5 markers, at least 6 markers, at least 7 markers, at least 8 markers, at least 9 markers, at least 10 markers, at least 11 markers, at least 12 markers, at least 13 markers, at least 14 markers, at least 15 markers, at least 16 markers selected from the group consisting of: CSFlPO, TPOX, VWA, D5S818, D7S820, D13S317, D3S1358, D16S539, D2S1338, A (gender marker Amelogenin), D8S1179, D21S11, D18S51, D19S433, THOl and FGA.
  • the STR markers comprise at least the group consisting of CSFlPO, FGA, THOl, TPOX, VWA, D3S1358, D5S818, D7S820, D8S1179, D13S317, D16S539, D18S51 and D21S1 l or at least the group consisting of D3S1358, VWA, D16S539, D2S1338, A (gender marker Amelogenin), D8S1179, D21S11, D18S51, D19S433, TH01 and FGA.
  • Extracting information from the biological sample may also comprise the use of a microarray.
  • a microarray allows parallel processing of a large number of markers and can be used for identification of markers in the form of snp's, STRs etc. The same microarray may be used to extract both the first and second information. Microarrays may also been applied as arrays in suspension.
  • Extracting first and second information from the biological sample may also comprise the use of methods relating to cytometry such as flow cytometry.
  • the invention may, in particular, find application in connection with human beings, but the invention is not limited to such application, as it may be foreseen that owners of pets, high-value racing horses or camels, high-value breeding animals (cows, sheep, etc.) may also desire a restriction on the flow of information related to their animals. Accordingly, subject may therefore refer to any animals however in a preferred embodiments subject refers to a human being (homo sapiens).
  • Bio sample comprise any biological material, which can be collected and subjected to extraction of at least a first information.
  • the biological sample may be matter obtained from microorganisms, plant, animals such as birds, fish and mammal.
  • the biological sample may also be a mixture of biological matter from different species, such a sample of meat infected with a microorganism such as bacteria.
  • the biological sample originate from a mammal such as a human being.
  • Body dust in the meaning of any biological material, which is disconnected
  • Body dust of mammals such as humans comprise saliva, blood and hair with some root cells, contain nucleus DNA and the hair it selves contains mitochondrial-DNA (mtDNA).
  • mtDNA mitochondrial-DNA
  • Body dust of humans may be collected at any place populated by humans such as in the house where one lives but basically at every place one has been like a hotel room, a toilet, a car, a taxi, the table in a restaurant and many other places, body dust may be found. All material lost by the body will be referred to as body- dust.
  • body dust such as body dust of humans may be regarded as a biological sample stored at conditions that favour degradation at least until it is "re-collected” and stored at conditions that do not favour degradation of biological material (e.g. in a buffer at low temperature).
  • the biological matter of the body dust sample is therefore in a state of degradation higher than when the matter where still a part of the body (the soma).
  • the first information QC-INF related to the quality of said biological sample 12 comprises
  • - information being extracted by analysis of genetic material comprising DNA or RNA or comprises - information being extracted by analysis of non-genetic biological material comprising the physical state or level of one or more proteins or polypeptides or peptides, or the biological state of cells or tissue, or any combination of the aforementioned.
  • the first information (QC-INF) related to the quality of said biological sample 12 is defined by the integrity of the biological matter from which the information is being extracted.
  • Integrity refers to the state functional of the biological matter.
  • DNA of a sample in a functional form organized in a chromosomal structure The integrity of the DNA may be severely affected if the organism comprising the DNA dies and the DNA is exposed to conditions that favor the hydrolysis of the DNA backbone. In the process of degradation from functional DNA to fragmented DNA the DNA loose integrity.
  • a polypeptide of a biological sample properly folded into a functional structure loose integrity when the polypeptide is exposed to proteolytic activity present in the sample.
  • cells of a biological sample may display a fully functional cell membrane recently after the collection of the sample. Gradually the cell membrane integrity is affected and the cells finally die from injures (e.g. caused by the nature of the sample buffer), necrosis, or apoptosis.
  • the genome consisting of DNA is for most organisms are arranged chromosomes.
  • the integrity of the genome of higher organisms are maintained by a complex system including system that repair damage caused to the genome and ensure that damages caused to the genome does not impair the process of replication.
  • Lower organisms such as bacteria have systems for maintaining the integrity of the genome, although these systems are less complex.
  • Cells comprised in a biological sample such as cells from a biopsy are not any longer in the natural context of the cells and will gradually loose integrity with a pace that depends upon the storage condition.
  • the cell may die by injury, necrosis or apoptosis and the DNA normally safely enveloped in the nucleus and mitochondria will be subject damaging conditions such as nucleases (endogenous as well as nucleases in the environment); humidity and acids causing hydrolysis; UV-light causing cross-linking.
  • nucleases endogenous as well as nucleases in the environment
  • humidity and acids causing hydrolysis
  • UV-light causing cross-linking If the sample is not stored at cold temperature the process of degradation will be significantly increased. Stains for DNA analysis must be thoroughly air-dried prior to sealing the paper pack-age to reduce the rate of degradation. Also storage at low temperatures (at 4 0 C or even better at -20 0 C) is beneficial.
  • the degree to which DNA is degraded in a sample dictates the size of DNA fragments that can be amplified, and thus the level of genetic information that can be retrieved. If a sample is too far degraded the remaining small fragments of DNA are of no use for extracting information such second information such as (DIAG-INF) according to the present invention. In contrast, a sample retaining high molecular weight DNA, regardless of age, can yield a wealth of data.
  • RNA is a notoriously fragile and labile, and fragments if the sample is stored incorrectly or for too long prior to performing the analysis extracting the biological information in question.
  • the extraction of information from a biological sample relying on RNA is even more sensitive to improper storage and/or too long storage.
  • the present system may be used to evaluate whether a sample such as a sample of qualify for the extraction of a second information, such as (DIAG-INF), related to the health and/or condition and/or identity of the subject (such as a mammal such as a human being).
  • a sample such as a sample of qualify for the extraction of a second information, such as (DIAG-INF), related to the health and/or condition and/or identity of the subject (such as a mammal such as a human being).
  • RNA quality including gel electrophoresis, Southern blot, gene-specific PCR, multiplex gene-specific PCR, short tandem repeat (STR) PCR and random amplified polymorphic DNA (RAPD) PCR. Although these methods provide information on the degree of DNA fragmentation, they may not all predict the ability of the sample to support the extraction of information from the sample by means of for example PCR.
  • Gel electrophoresis provide information at the level of fragmentation (e.g Figure 2). High molecular DNA representing DNA that is not fragmented will appear at the top of the gel. Degraded DNA will appear as a smear through the lane on the gel accumulating at the bottom of the gel. A ladder like structure may appear reflecting initial fragmentation of the DNA that is not protected by the histones in nucleosome structures.
  • Southern blot provides information at the level of DNA fragmentation only. Although, Southern blotting is more laborious than gel electrophoresis it requires less DNA. More advanced forms of electrophoresis are available such as capillary electrophoresis with low sample consumption.
  • Devices such as 96-channel microfabricated capillary array electrophoresis may be used to reduce sample consumption increase the throughput of samples.
  • electrophoresis nor Southern blot provide information at the level nucleotide cross-linking and nicks in the DNA that affect the performance of a PCR application.
  • Methods of PCR PCR can be used to determine the largest size class of DNA existing in each sample by targeting a series of amplification products. Progressively larger segments of DNA can be amplified until a negative result is obtained, this cut-off size can indicate how degraded a sample is. Amplicon sizes can then be compared within a weathering stage to see what generalities can be observed, within a stage and among stages, to discern any statistically meaningful differences in degradation levels.
  • the first information QC-INF related to the quality of said biological sample 12 comprises information being extracted by analysis of genetic material comprising DNA or RNA, wherein said quality is defined by the integrity of genetic material from which the information is being extracted.
  • the integrity of the genetic material is defined by the maximum obtainable amplicon size in a method of PCR. Accordingly, the maximum obtainable amplicon size in a method of PCR is a quality parameter QC-P of the biological sample and a specific maximum obtainable amplicon size is a threshold.
  • a biological sample that does not qualify the generation the products of a PCR equal to or above the size defined by the threshold parameter to the specified maximum obtainable amplicon size as threshold parameter for the first information QC-INF will not prompt an authorization flag AF and affect the extraction and/or processing of the second information DIAG-INF wherein the processing of the second information DIAG-INF into third information RES is dependent on the prompting of the authorization flag AF of said biological sample 12, and/or wherein accessing the second information DIAG-INF and/or accessing the third information RES is dependent on the prompting of the authorization flag AF of said biological sample 12, and/or wherein the execution of the analysis performed by the analyzing unit AU for the extraction of the second information DIAG-INF is dependent on the prompting of the authorization flag AF of said biological sample 12.
  • the size of the PCR products may refer to number of base pairs (bp). Alternatively, the size of the PCR product may refer to the mass of the DNA usually expressed in Daltons.
  • Gene-specific PCR as means of directly measures the properties of the sample in terms of amplifying a target DNA of a specific size using a specific primer set.
  • the method usually requires a minimum of 2 to 5 ng of DNA.
  • Cross-linking and nicks in the DNA is scattered across the entire genome and may affect the performance of the gene- specific PCR and indicate how well the sample may perform further PCR application such as a diagnostic method based on PCR.
  • the power of the assessment of the DNA quality of a sample may be increased by employing multiplex PCR typically on two to three DNA targets in a single PCR reaction.
  • the first information QC-INF being related to the quality of said biological sample 12, the first information being extracted by a method of quality control analysis, wherein said method of quality control analysis uses gene-specific PCR.
  • the first information QC-INF being related to the quality of said biological sample 12, the first information being extracted by a method of quality control analysis, wherein said method of quality control analysis uses gene-specific PCR.
  • INF being related to the quality of said biological sample 12
  • the first information being extracted by a method of quality control analysis, wherein said method of quality control analysis uses short tandem repeat DNA typing. If the DNA of a biological sample is degraded, its length can be reduced to a size range smaller than this, resulting in allelic or locus dropout and an incomplete genetic profile.
  • the integrity of the genetic material relates to the predicted size of the products of a STR PCR.
  • the predicted size of the products of a STR PCR is a quality parameter QC-P of the biological sample and a range around the predicted size of the products of a STR PCR is a threshold.
  • a biological sample that does not qualify to the generation the products of a STR PCR within the specified range defined by the threshold parameter for the first information QC-INF will not prompt an authorization flag AF and affect the extraction and/or processing of the second information DIAG-INF wherein the processing of the second information DIAG-INF into third information RES is dependent on the prompting of the authorization flag AF of said biological sample 12, and/or wherein accessing the second information DIAG-INF and/or accessing the third information RES is dependent on the prompting of the authorization flag AF of said biological sample 12 , and/or wherein the execution of the analysis performed by the analyzing unit AU for the extraction of the second information DIAG-INF is dependent on the prompting of the authorization flag AF of said biological sample 12.
  • the size of the STR PCR products may refer to number of base pairs (bp).
  • Randomly amplified polymorphic DNA (RAPD) PCR generates DNA fragments with a broad size range.
  • the range of the product size reflects the degree of DNA fragmentation of the sample in an unbiased manner.
  • the method requires a little as 2 ng of
  • the corresponding sets of quality control parameters QC-P relating to the first information QC-INF refers to an control (run internal or parallel control sample), and the threshold parameter for prompting said authorization flag AF defined by the products of the control number of products.
  • An authorization flag AF for said biological sample 12 is prompted if the extraction of the first information relating to number of products of a PCR is within the specified range of the control.
  • an authorization flag AF for said biological sample 12 is prompted if the number of products generated from said biological sample 12 is more than 50% of the corresponding products of the control, for example more as 75% of the products of the control, such as more as 85% of the products of the control, for example more as 90 % of the products of the control, such as more as 95 % of the products of the control, for example 100 % of the products of the control.
  • the corresponding sets of quality control parameters QC-P relating to the first information QC-INF refers to a control (run internal or parallel control sample), and the threshold parameter for prompting said authorization flag AF defined by the products of the control such as size.
  • An authorization flag AF for said biological sample 12 is prompted if the extraction of the first information relating to size of products of a PCR is within the specified range of the control.
  • an authorization flag AF for said biological sample 12 is prompted if the size of products generated from said biological sample 12 is within a 50% base pair range of the corresponding products of the control, for example within a 75% base pair range of the products of the control, such as within a 85% base pair range of the products of the control, for example within a 90% base pair range of the products of the control, such as within a 95% base pair range of the products of the control, for example match the size of the products of the control.
  • the corresponding sets of quality control parameters (QC-P) relating to the first information (QC-INF) refers to a ratio between the quantity of at least to products generated by the method of analysis such as PCR, and the threshold parameter for prompting said authorization flag AF defined by preset ratio between the quantity of at least to products.
  • An authorization flag AF for said biological sample 12 is prompted if the extraction of the first information relating a ratio between the quantity of at least two products generated by the method of analysis in within a specified range of the threshold parameter.
  • the threshold parameter for prompting said authorization flag AF defined by a preset ratio between the quantities of at least to products, wherein the first product is generated from amplification of the locus D19S433 marker and the second product is generated from amplification of the locus D18S15 marker.
  • the threshold parameter for prompting said authorization flag AF set to a D19S433/D18S15 ratio (D 19/Dl 8 ratio). The method is further detailed in Example 1.
  • the method of assessing the quality of the DNA of a sample should provide the necessary information needed to evaluate whether the sample qualifies for further extraction of information such as a PCR method for the purpose of diagnostic.
  • the DNA quality of a sample should not be assessed using gel electrophoresis if the sample subsequently used for extraction of further information means of method, which are sensitive to cross-linking and nicks in the DNA.
  • the system wherein the first information QC-INF related to the quality of said biological sample 12 is defined by the integrity of the biological matter from which the information is being extracted, said integrity of said genetic material is defined by the state of degradation of said genetic material.
  • the genetic material comprises DNA (nuclear and mitochondrial DNA) and RNA.
  • the degradation of the genetic material may be assessed by any of methods for assessing DNA quality know in the art including the methods mentioned herein. Since RNA is less resistant to degradation than DNA and thus more rapidly degraded from the sample, the degree of DNA degradation in the biological sample may be used as indicator of RNA degradation as well. Alternatively, the quality of the RNA in the sample may be assessed using a method suitable for the purpose.
  • the extraction of the first information QC-INF from the biological sample 12 by a method of analysis comprises use of nucleotide sequence amplification, a microarray, cytometry, or electrophoresis.
  • the methods comprise the use of a least one method, for example a nucleotide sequence amplification step and extraction of the information from the nucleotide sequence amplification by means of electrophoresis.
  • the analyzing unit according to the invention is capable of performing at least one method in order to extract the first information QC-INF from the biological sample 12.
  • Sample quality assessment to evaluate whether the biological sample qualifies for the extraction of the second information according to the invention may require the extraction of the first information from biological matter other than the genetic matter of the biological sample.
  • first information relating to the total or relative cell count of a subpopulation of cell in a sample may more appropriate if the second information relates the extraction of information regarding the presence of a specific surface receptor on said subpopulation of cells.
  • the extraction of the information according to the invention may use cytometric methods such as using retention of or exclusion of fluorescent dyes (e.g propidium iodide and fluorescein) as markers in cell membrane integrity relating to the first information QC-INF; and an anti-body for the identification of the subpopulation of cells as marker relating to the second information.
  • fluorescent dyes e.g propidium iodide and fluorescein
  • the first information according to the invention comprises the use of any markers such as dye, anti-bodies and other probes related to the integrity of cells or tissue comprised in the sample.
  • Diagnostic methods based on analysis of other material than DNA may be even more sensitive to degradation. Analysis of blood samples by means of cytometry most often rely on fresh samples. Stored at room temperature and/or in solution not appropriate for storage, the cells rapidly loose membrane integrity, die and disintegrate by the process of apoptosis or necrosis. Rare population of cells subject for analysis may be eliminated from the sample. Subjecting the sample to analysis such as a diagnostic assay therefore may apart from wasting time and costly reagent and even result in the wrong conclusion.
  • the first information relates to cell membrane integrity of the cells in said biological sample.
  • the integrity cells or tissue of the sample is defined by the state of cyto logical markers for cellular integrity comprising markers for cell membrane integrity, apoptosis and necrosis.
  • the appropriate evaluation of the first information may at the level of proteins, polypeptides or peptides reflect the state of protein degradation in the sample.
  • the first information relates to the integrity of the at least one protein, polypeptide or peptide of the biological sample.
  • the integrity of the proteins or polypeptides or peptides of the biological sample is defined by the state of degradation or posttranslational modification associated with degradation of said proteins or polypeptides or peptides.
  • PCR multiplex kits for STRs like the PowerPlex® 16 kit from Promega Corporation or the AmpFISRT® IdentifilerTM kit from Applied Bio systems are suited, however some optimization is possible, e.g.:
  • loci Fewer loci can be used, and loci don't have to show high variability, because there is no need for identification.
  • the set of loci covers a wide range of sizes.
  • a single type of fluorescence label is sufficient under the condition that all amplicons sizes are distinct (electrophoreses). If instead of the sizes the actual sequences (or part of it) of amplicons are used for identification (microarrays), their sizes do necessarily not have to be distinct.
  • the second information preferably relates to the health and/or condition and/or identity verification of the subject (such as a mammal e.g. a human being) from which the sample was obtained.
  • the second information DIAG-INF related to the health and/or condition and/or identity verification of said subject 10 comprises
  • non-genetic biological material comprising the physical state or level of one or more proteins or polypeptides or peptides, or the biological state of cells or tissue, or any combination of the aforementioned.
  • Second information relating to the health and/or condition of the extracted from biological sample of a subject is often used for purpose of predicting the risk of having a specific disease.
  • the first information QC-INF and the second information DIAG-INF comprise overlapping information.
  • the system of the present invention may be used prevent abuse of body-dust for DNA-analysis.
  • the body-dust left at any place may contain full-fledged DNA applicable to a wide range of applications in the field of DNA profiling. This implies that any third party can use DNA left in body-dust for identification and diagnostic purposes without approval from the person from which the sample originate.
  • Today many services are provided to do identity and/or family-relationship testing. With the emergence of very advanced biosensors such testing becomes very cheap and easy to perform. Even in the home situation such tests can be performed.
  • Another example is body-dust of animals such as a race horse, where any DNA profiling for whatever reason may require the approval of the owner of the animal.
  • Biosensors and diagnostic systems are getting more advanced and in the future it may be possible to acquire and use the system on biological sample for applications such as diagnostics.
  • the system according to the present invention may prevent the use of biosensors and diagnostic systems for the purpose of analyzing body dust.
  • the system further provides an intelligent management of the consumption of reagents.
  • Reagent consumption management may be of particular relevance if the system is installed and used on location, which are not certified laboratories and/or handled by personnel with no certified training in the of lab equipments. Examples of such system are handheld biosensors for self-operation and system installed in remote areas for self- operation. Thus, the system may extend the operational times-span between services such as refilling the system with reagents.
  • FIG. 7 A flow chart of a method according to the present invention is shown in Fig. 7. The method comprises:
  • the first information QC-INF being related to the quality of said biological sample 12, the first information being extracted by a method of quality control analysis, and b) the second information DIAG-INF being related to the health and/or condition and/or identity verification of said subject 10,
  • the quality control database QC-DB arranged to receive the first information QC-INF from the analyzing unit AU, the quality control database comprising corresponding sets of quality control parameters QC-P, an authorization flag AF for said biological sample 12, and a threshold parameter for prompting said authorization flag AF, and
  • Example 1 Various methods are developed to assess the degree of degradation using standard STR kits.
  • a first method is the use of standard probes in a kit for measuring the quantity of 2 loci and to determine their ratio.
  • the AmpFISRT® Identif ⁇ lerTM kit from Applied Biosystems is used to determine the presence of D19S433 (about 120 bp) and D18S15 (about 300 bp).
  • the D 19/D 18 ratio is determined (table 1 , columns 6 and 7).
  • qPCR quantitative PCR
  • the data of the D19/D18 ratio is sorted and a threshold is given at 1.
  • the control DNA (sample 23) has the lowest score and a single blood (sample 3) and buccal swap (sample 22) are close over the threshold. Many of the samples are far beyond threshold and can therefore easily be considered as degraded.
  • the threshold for deciding whether a sample is degraded or not can be set to optimize the error probabilities like the false acceptance ration (FAR) and the false rejection ratio (FRR).
  • the BRCAl and BRC A2 proteins are tumor suppressors involved in DNA repair and play a critical role in embryonic development. Mutations in the BRCAl and
  • BRC A2 genes are associated with increased risk of developing breast and ovarian cancers (BOC). More than 600 of mutations of which many are associated with increased risk of BOC have been identified in the BRACl gene alone. Subjects at risk may be identified by diagnostic methods such as methods based on PCR and sequencing, Multiplex Ligation-dependent Probe Amplification (MLPA), Quantitative Multiplex PCR of Shorts Fluorescents Fragments (QMPSF), heteroduplex analysis (HDA) by multi- capillary electrophoresis or oligonucleotide array based on comparative genomic hybridization (array-CGH).
  • MLPA Multiplex Ligation-dependent Probe Amplification
  • QMPSF Quantitative Multiplex PCR of Shorts Fluorescents Fragments
  • HDA heteroduplex analysis by multi- capillary electrophoresis or oligonucleotide array based on comparative genomic hybridization (array-CGH).
  • Clinical practice guidelines recommends that women whose family history is associated with an increased risk for deleterious mutations in BRCAl or BRCA2 genes be referred for genetic counseling and evaluation for BRCA testing.
  • the guidelines recommends against routine referral for genetic counseling or routine breast cancer susceptibility gene (BRCA) testing for women whose family history is not associated with an increased risk for deleterious mutations in breast cancer susceptibility gene 1 (BRCAl) or breast cancer susceptibility gene 2 (BRCA2).
  • Remedies for risk reduction are available, the approaches are drastic.
  • Female carriers of BRCAl and BRC A2 mutation associated with increased risk of BOC are frequently subjected to prophylactic oophorectomy and/or mastectomy. Women at risk of breast cancer, but currently well, occasionally choose to have such drastic prophylactic surgery.
  • the system of the present invention may prevent insurers and employers from using such predictive genetic testing without authorization to obtain a genetic profile of subject (in this case on the BRCAl and BRCA2 genes) based on a biological sample such as body dust. Such information may be used to evaluate who gets insurance or a job.
  • the system of the present invention may be employed to ensure that the second information DIAG-INF in this case the genetic profile related to the BRCA mutation testing rely on a sample comprising DNA, which are not degraded.
  • the system may therefore prevent any third party from submitting a biological sample obtained from another person to said BRCA testing and acquire delicate information about another person.
  • the system is adapted for extracting at the first QC-INF information, which in this case is the DNA quality of the sample.
  • the first information relates to the predicted size of the products of a STR PCR.
  • the first information is transmitted to the quality control database QC-DB arranged to receive the first information QC-INF from the analyzing unit AU extracting the information from the sample, the quality control database comprising corresponding sets of quality control parameters QC-P related to size of the generated PCR product relative to the predicted size of the products, an authorization flag AF for said sample, and a threshold parameter for prompting said authorization flag AF.
  • the predicted size of the products of a STR PCR is a quality parameter QC-P of the sample and a range around the predicted size of the products of a STR PCR is a threshold. If the sample does not match the specs for the good quality DNA in the meaning of generation products within a predefined range of the predicted size of the products of a STR PCR, further extraction of information, the second information DIAG-INF is rejected.
  • the second information in this case relates to the presence or absence of specific mutation(s) in BRCAl or BRC A2 gene of the DNA obtained from the subject.
  • the second information may be extracted from the sample by means of PCR using specific primers designed to detect the mutations in question.
  • the second information may be extracted by means of oligonucleotide array based on comparative genomic hybridization (array-CGH) as described recently in Rouleau-E, et al. (2007)
  • the second information is transmitted to the processing unit PU arranged for receiving the second information DIAG-INF from the analyzing unit AU, which processes the second information into third information RES.
  • the invention can be implemented by means of hardware, software, firmware or any combination of these.
  • the invention or some of the features thereof can also be implemented as software running on one or more data processors and/or digital signal processors.
  • the individual elements of an embodiment of the invention may be physically, functionally and logically implemented in any suitable way such as in a single unit, in a plurality of units or as part of separate functional units.
  • the invention may be implemented in a single unit, or be both physically and functionally distributed between different units and processors.

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Abstract

L'invention concerne un système permettant de traiter des informations obtenues à partir d'un échantillon biologique d'un sujet. Le système comporte une unité d'analyse (AU) destinée à analyser les informations pouvant être obtenues à partir de l'échantillon biologique. L'unité d'analyse est adaptée pour extraire au moins une première information (QC-INF) et une deuxième information (DIAG-INF) à partir dudit échantillon biologique. La première information (QC-INF) concernant la qualité de l'échantillon biologique est extraite par un procédé d'analyse de contrôle qualité et la deuxième information (DIAG-INF) concerne la vérification de la santé et/ou l'état et/ou l'identité dudit sujet. L'exécution de l'analyse, du traitement de ou de l'accès à la deuxième information (DIAG-INF) en une troisième information (RES) dépend du de l'indicateur d'autorisation (AF) dudit échantillon biologique (12). Le système donne des moyens permettant d'éviter l'application d'échantillons prélevés chez des sujets sans leur connaissance et approbation, tels que des poussières corporelles (par ex. un cheveu, des tâches de sang).
PCT/IB2009/052553 2008-06-20 2009-06-16 Système et procédé permettant d'évaluer la qualité d'un échantillon WO2009153731A1 (fr)

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN111933255A (zh) * 2020-06-17 2020-11-13 戴纳智慧医疗科技有限公司 一种对样本转送的审批管理方法

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WO2007053491A2 (fr) * 2005-10-28 2007-05-10 The Trustees Of Columbia University In The City Of New York Procede et trousse d'evaluation de la qualite d'un arn
WO2007064294A1 (fr) * 2005-11-29 2007-06-07 Denator Aktiebolag Procede de determination de la qualite d’un echantillon biologique

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Publication number Priority date Publication date Assignee Title
WO2007053491A2 (fr) * 2005-10-28 2007-05-10 The Trustees Of Columbia University In The City Of New York Procede et trousse d'evaluation de la qualite d'un arn
WO2007064294A1 (fr) * 2005-11-29 2007-06-07 Denator Aktiebolag Procede de determination de la qualite d’un echantillon biologique

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN111933255A (zh) * 2020-06-17 2020-11-13 戴纳智慧医疗科技有限公司 一种对样本转送的审批管理方法

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