US3716632A - Test composition and method for the in vitro determination of thyroid function - Google Patents

Test composition and method for the in vitro determination of thyroid function Download PDF

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Publication number
US3716632A
US3716632A US00032396A US3716632DA US3716632A US 3716632 A US3716632 A US 3716632A US 00032396 A US00032396 A US 00032396A US 3716632D A US3716632D A US 3716632DA US 3716632 A US3716632 A US 3716632A
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Prior art keywords
indicator
composition
blue
thyroid
test composition
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Expired - Lifetime
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US00032396A
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English (en)
Inventor
M Fader
J Christner
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Bayer Corp
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Miles Laboratories Inc
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    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/74Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving hormones or other non-cytokine intercellular protein regulatory factors such as growth factors, including receptors to hormones and growth factors
    • G01N33/78Thyroid gland hormones, e.g. T3, T4, TBH, TBG or their receptors
    • 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
    • Y10S436/00Chemistry: analytical and immunological testing
    • Y10S436/804Radioisotope, e.g. radioimmunoassay
    • 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
    • Y10S436/00Chemistry: analytical and immunological testing
    • Y10S436/826Additives, e.g. buffers, diluents, preservatives

Definitions

  • thyroxine-binding proteins are the primary binding sites for the hormones produced by the thyroid gland. Since the TBF have a strong affinity for thyroid hormone, they will bind any free thyroid hormone available until they become saturated. In this test, such hormone is made available to the TB? in a sample of the-patients serum and a secondary binding site consisting of a resin powder or crosslinked dextran (Sephadex) is provided to take up any of the added radioactive triiodothyronine or thyroxine (hereinafter referred to as T or T respectively) not bound by the TBP.
  • T or T radioactive triiodothyronine
  • the main object of the present invention is to provide a thyroid function test solution containing a pH indicator which possesses the three fold advantage of (1) providing a color change when the serum sample is added thereto (2) inhibiting the formation of free iodide from radioactive iodinated T and (3) decreasing the adsorption of radioactive iodinated T to the walls of the vessel containing the same.
  • thyroid function tests may be performed in many different ways, one common method involves adding an appropriate amount ofa buffered aqueous solu tion of radioactive triiodothyronine or thyroxine and serum to a column of ion exchange resin such as Amberlite IRA 400 made by Rohm and Haas Co. of Philadelphia, Pa. or a cross linked dextran such as Sephadex G-25 made by Pharmacia Corp. of Uppsala, Sweden.
  • the column is then drained and the radioactivity measured in a gamma ray counting instrument for 1 minute.
  • the column is thereafter eluted with distilled water and the radioactivity remaining on the column is again measured for 1 minute.
  • the percent retention is then calculated by comparing the two readings. Retentions of 41 percent to 6l percent are considered normal with cross-linked dextran (Sephadex) columns and 25 percent to 35 percent with resin columns.
  • the pH indicator employed in the present invention is of the protein error type which changes color in the presence of albumin without a significant actual change in pH.
  • an indicator is added in a concentration of about 5. to micrograms per milliliter of aqueous I -T reagent buffered at a pH of 5 the reagent changes from its initial color to a second color when the serum is added.
  • This second color acts as a marker which will appear at varying heights on the resin or dextrancolumndepending on the last operation step completed in the test procedure, since the color will move with the solvent front.
  • a preferred indicator is bromcresol purple which changes color from yellow to purple within the pH range of 5.2 to 6.8.
  • the yellow-orange color will turn to a grey-green color when serum is added at a ratio of 0.05 milliliter of serum to 0.45 milliliter of reagent mixture.
  • the green color indicates the presence of the serum.
  • the improved T test reagent with pH indicator of this invention will contain the following:
  • a buffered aqueous solution of T containing 60 micrograms of bromcresol purple per milliliter prepared as shown in the foregoing example was stored under various conditions in a polyethylene container and the percent of free iodide formed was compared with a similar solution containing no pH indicator.
  • the initial solutions employed contained about 2 percent free iodide.
  • the radioactive T solution containing the indicator showed 2.25 percent free iodide compared to 5.2 percent free iodide in the control sample containing no indicator.
  • the T test solution with pH indicator contained 4.25 percent free iodide, whereas the control sample had a free iodide content of 9.0 percent.
  • a still further advantage of the use of a pH indicator in T test solution is to inhibit the adsorption of radioactive T to the walls of a plastic or glass container.
  • Propylene glycol and albumin have been used in the past to reduce such adsorption but these additives interfere with the thyroid function test.
  • aqueous radioactive T solutions containing one of several indicators at a concentration of 60 micrograms per milliliter of solution were rotated on a wheel in a polyethylene container for 14 days and the percent activity remaining was compared with a control sample containing no indicator.
  • prewashed means contacting (such as by rinsing, soaking or flushing) the interior surfaces of said containers with an aqueous, buffered solution of one of the pH indicators hereinbefore disclosed.
  • Bromcresol purple or bromthymol blue are the preferred indicators. These are employed at a concentration of from 50 to 70 micrograms per milliliter of water buffered to a pH of about 5.
  • T employed in the test compositions of this invention can be made radioactive with either F or 1", although l is preferred, since it has a half-life of 56 days compared to 1"" which has a half-life of 8 days.
  • a test composition for determining thyroid function comprising a radioactive thyroid hormone and a buffer for establishing the pH of thecomposition in a predetermined range
  • the improvement which comprises adding to the composition a color responsive is used to maintain the pH of the composition at about 5 3.
  • the indicator is selected from the group consistin of bromcresol purple, chlorphenol red, tetrabromp enol blue,
  • bromphenol blue bromcresol green and bromthymol blue.
  • test composition as in claim 1 wherein the indicator is bromcresol purple.
  • a test composition as in claim 1 wherein the indicator is bromthymol blue.
  • a test composition as in claim 1 wherein the thyroid substance is triiodothyronine.
  • a method as in claim 8 wherein the indicator is selected from the group consisting of bromcresol purple, chlorphenol red, tetrabromphenol blue, bromphenol blue, bromcresol green and bromthymol blue.
  • a method as in claim 8 wherein the thyroid substance is triiodothyronine.
  • a method of inhibiting the adsorption of radioactive triiodothyronine from an aqueous solution thereof on the walls of a polyethylene container which comprises adding to said aqueous solution an adsorption inhibiting amount of a protein error type pH indicator.
  • a method as in claim 13 wherein the indicator is selected from the group consisting of bromcresol purple, chlorphenol red, tetrabromphenol blue, bromphenol blue, bromcresol green and bromthymol blue.

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Molecular Biology (AREA)
  • Engineering & Computer Science (AREA)
  • Endocrinology (AREA)
  • Biomedical Technology (AREA)
  • Chemical & Material Sciences (AREA)
  • Urology & Nephrology (AREA)
  • Hematology (AREA)
  • Immunology (AREA)
  • Microbiology (AREA)
  • Cell Biology (AREA)
  • Biotechnology (AREA)
  • Food Science & Technology (AREA)
  • Medicinal Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Investigating Or Analysing Biological Materials (AREA)
US00032396A 1970-04-27 1970-04-27 Test composition and method for the in vitro determination of thyroid function Expired - Lifetime US3716632A (en)

Applications Claiming Priority (1)

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US3239670A 1970-04-27 1970-04-27

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US3716632A true US3716632A (en) 1973-02-13

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US (1) US3716632A (OSRAM)
CA (1) CA956097A (OSRAM)
DE (1) DE2120442B2 (OSRAM)
FR (1) FR2090714A5 (OSRAM)
GB (1) GB1347498A (OSRAM)

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3960492A (en) * 1974-05-31 1976-06-01 Nuclear Diagnostics, Inc. Method for determining an index of binding protein content of blood
US3985867A (en) * 1974-01-02 1976-10-12 The Radiochemical Centre Limited Immunoassays employing a colored second antibody
US4108974A (en) * 1976-08-27 1978-08-22 Bio-Rad Laboratories, Inc. Radioimmunoassay for thyroid hormone
US4278653A (en) * 1979-02-01 1981-07-14 New England Nuclear Corporation Methods and kits for double antibody immunoassay providing a colored pellet for easy visualization
US4299812A (en) * 1978-11-29 1981-11-10 Diagnostic Products Corp. Immunoassay of thyroxine in neonates using dried blood samples
US4311690A (en) * 1978-06-20 1982-01-19 Damon Corporation Test set and method for the determination of free hormones

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3485587A (en) * 1956-02-29 1969-12-23 Miles Lab Protein indicator
US3507618A (en) * 1964-11-27 1970-04-21 Squibb & Sons Inc Apparatus and method for determining thyroid function
US3516794A (en) * 1964-12-14 1970-06-23 Squibb & Sons Inc Apparatus and method for determining thyroid function

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3485587A (en) * 1956-02-29 1969-12-23 Miles Lab Protein indicator
US3507618A (en) * 1964-11-27 1970-04-21 Squibb & Sons Inc Apparatus and method for determining thyroid function
US3516794A (en) * 1964-12-14 1970-06-23 Squibb & Sons Inc Apparatus and method for determining thyroid function

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3985867A (en) * 1974-01-02 1976-10-12 The Radiochemical Centre Limited Immunoassays employing a colored second antibody
US3960492A (en) * 1974-05-31 1976-06-01 Nuclear Diagnostics, Inc. Method for determining an index of binding protein content of blood
US4108974A (en) * 1976-08-27 1978-08-22 Bio-Rad Laboratories, Inc. Radioimmunoassay for thyroid hormone
US4311690A (en) * 1978-06-20 1982-01-19 Damon Corporation Test set and method for the determination of free hormones
US4299812A (en) * 1978-11-29 1981-11-10 Diagnostic Products Corp. Immunoassay of thyroxine in neonates using dried blood samples
US4278653A (en) * 1979-02-01 1981-07-14 New England Nuclear Corporation Methods and kits for double antibody immunoassay providing a colored pellet for easy visualization

Also Published As

Publication number Publication date
DE2120442A1 (de) 1971-11-11
GB1347498A (en) 1974-02-27
DE2120442C3 (OSRAM) 1973-11-29
FR2090714A5 (OSRAM) 1972-01-14
CA956097A (en) 1974-10-15
DE2120442B2 (de) 1973-05-10

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