Immunochemical methods for the detection of antibody against HTLV-III

• US 4,774,175 A
• Filed: 03/01/1985
• Issued: 09/27/1988
• Est. Priority Date: 03/01/1985
• Status: Expired due to Term

• Current Assignee: Centocor Incorporated (Johnson & Johnson), Repligen Corporation
• Original Assignee: Centocor Incorporated (Johnson & Johnson)
• Inventors: Kato, Ikunoshin, Chang, Nancy T., Chang, Tse W., Chanda, Pranab
• Primary Examiner(s): Nucker, Christine M.
• Application Number:
  US06/707,066
• Time in Patent Office: 1,306 Days
• Field of Search: 435/5, 435/7, 435/180, 435/805, 530/324, 436/518, 436/528, 436/531, 436/811
• US Class Current: 435/5
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: C07K 14/005 from viruses
• CPC Class Codes: C12N 2740/16122 New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
• CPC Class Codes: C12Q 1/703 Viruses associated with AIDS
• CPC Class Codes: Y10S 435/805 Test papers
• CPC Class Codes: Y10S 435/974 Aids related test
• CPC Class Codes: Y10S 436/811 Test for named disease, body condition or organ function
• CPC Class Codes: Y10S 930/221 Retrovirus related, or human immunodeficiency virus related, or simian immunodeficiency virus related

Abstract

Gene segments of human T cell lymphotropic virus type III (HTLV-III) were expressed in E. coli as peptides that are reactive with sera from patients with acquired immune deficiency syndrome (AIDS). Among recombinant peptides one designated HTLV-III polypeptide 121, contained 85 amino acid residues encoded by a gene segment in the env-lor region of the HTLV-III genome. The polypeptide is strongly reactive with AIDS patient sera. The peptide produced and purified as a fusion protein on a large scale. Solid phase immunoassays employing this recombinant peptide as an immunoabsorbent can reliably and reproducibly detect antibodies in sera of patients with HTLV-III infection. In two representative serum panels, the assay detected the presence of antibodies in 120 of 121 sera from patients with AIDS or AIDS-related complex (ARC), and only in 1 of 92 normal controls. Based upon HTLV-III polypeptide 121 as immunoreactive agent, sensitive and specific immunoassays for HTLV-III infection have been developed.

Claims

1. An immunoreactive HTLV-III derived polypeptide comprising HTLV-III polypeptide 121 having the sequence ##STR4##

2. In an immunochemical assay for detecting or quantifying antibody against HTLV-III in a fluid comprising contacting the fluid with an immunoreactive HTLV-III-derived antigen under conditions which allow the antigen to bind anti-HTLV-III antibody in the fluid, the improvement wherein the immunoreactive HTLV-III-derived polypeptide comprises HTLV-III polypeptide 121 having the sequence ##STR5##

3. A method of detecting antibody against HTLV-III in a biologial fluid, comprising the steps of:

(a) incubating an immunoadsorbent comprising a solid phase to which is attached an immunoreactive HTLV-III derived polypeptide comprising HTLV-III polypeptide 121 having the amino acid sequence ##STR6## with a sample of the biological fluid to be tested, under conditions which allow anti-HTLV-III antibody in the sample to bind to the immunoadsorbent;

(b) separating the immunoadsorbent from the sample; and

(c) determining if antibody has bound to the immunoadsorbent as an indication of anti-HTLV-III in the sample.

4. A method of claim 3, wherein the step (c) of determining if antibody has bound to the immunoadsorbent comprises incubating the immunoadsorbent with a labeled antibody against antibody of the species from which the biological fluid is derived;

separating the immunoadsorbent from the labeled antibody after the incubation period; and

detecting the label associated with the immunoadsorbent as an indication of antibody bound to the immunoadsorbent.

5. A method of claim 3, wherein the step c of determining if antibody has bound to the immunoadsorbent comprises incubating the immunoadsorbent with laeled HTLV-III polypeptide 121;

separating the immunoadsorbent from the labeled HTLV-III polypeptide 121; and

detecting the label associated with the immunoadsorbent as an indication of antibody bound to the immunoadsorbent.

6. A method of claim 3, wherein the step c of determining if antibody has bound to the immunoadsorbent comprises incubating the immunoadsorbent with labeled Proten A;

separating the immunoadsorbent from the labeled Protein A; and

detecting the label associated with the immunoadsorbent as an indication of antibody bound to the immunoadsorbent.

7. A method of detecting antibody against HTLV-III in a human serum or plasma sample, comprising the steps of:

(a) providing an immunoadsorbent comprising a bead coated with an immunoreactive HTLV-III derived polypeptide comprising of HTLV-III polypeptide 121 having the amino acid sequence ##STR7## with a sample of the human serum or plasma sample to be tested, under conditions which allow anti-HTLV antibody in the sample to bind to the immunoadsorbent;

(b) separating the immunoadsorbent from the sample; and

(c) determining if antibody has bound to the immunoadsorbent as an indication of anti-HTLV-III in the sample.

8. A method of claim 7, wherein the immunoabsorbent further comprises a coat of animal protein.

9. A method of claim 7, wherein the labeled anti-(human IgG) antibody is an animal antibody and the serum or plasma sample is diluted with normal serum of an animal of the same species.

10. A method of claim 9, wherein the anti-(human IgG) antibody is a goat antibody and the serum or plasma sample is diluted with normal goat serum.

11. A method of claim 7, wherein the anti-(human IgG) antibody is labeled with a radioisotope, an enzyme or a fluorescent compound.

12. An immunoadsorbent for use in a solid phase immunochemical assay for antibody against HTLV-III, comprising a solid phase to which is affixed an immunoreactive HTLV-III polypeptide comprising polypeptide 121 having the sequence:

##STR8##

13. An immunoadsorbent of claim 12, wherein the solid phase is a glass or plastic bead, a well of a microtiter plate or a test tube.

14. An immunoadsorbent of claim 12 further comprising a post-coat of animal protein.

15. A substantially immunologially pure preparation of HTLV-III polypeptide 121 having the amino acid sequence ##STR9##