A PEPTIDE COMPOUND AND ANTISERUM PRODUCED BY USING THIS COMPOUND

Abstract: Peptide compounds containing amino-acid residues occuring naturally in amyloid A protein and/or serum amyloid A protein and specially intended for the production of specific antisera for amyloid A protein and/or serum amyloid A protein. It is the characteristic feature of the peptide compounds that the sum of the hydrophilicity values of the respective amino-acid residues is greater than, or equal to, 0. Specific antisera for amyloid A protein and/or serum amyloid A protein and produced using a peptide compound of the type specified above.

Description: TECHNICAL FIELD Peptide compound and antisera, produced by the use of such compound, will be described. The invention relates in particular to peptide compounds for the production of antisera which are specific for amyloid A protein and/or serum amyloid A protein (hereafter called AA and SAA respectively) and which consequently constitute valuable means for the detection and the determination of the concentration of the said proteins. BACKGROUND ART In many chronic illnesses, such as rheumatoid arthritis, tuberculosis, hepatitis and infectious and tumerous affections there is a strong increase in the blood content of SAA. Through the splitting off of the 28 last amino-acid residues from SAA in such affection AA is formed in organs of these patients to such an extent that they cease to function. The organs which are hit earliest are the kidneys and the heart. This illness manifestation is called amyloidosis and, if not treated, is lethal. To make it possible to detect amyloidosis at an early stage and thus allow treatment of this illness manifestation through increased action against the basic affection, a method is required for the detection of AA in tissue specimens and/or the determination of the SAA content in blood samples. A known method which is based on the production of antisera which are specific for AA and/or SAA is described in the British Medical Journal, vol. 288, 4th February 1984. As far as we know this known method has not become generally accessible since it has proved particularly difficult to produce antisera specific for AA and/or SAA. Moreover, the method has been subject of certain criticism (see British Medical Journal, vol. 288, 17th March 1984, pp. 859-60. It is an object of the invention, therefore, to overcome the difficulty of the known technique and thereby lay the foundation of a well-functioning method of the type described above. The object is achieved in accordance with the invention with the help of the peptide compunds and antisera which are defined in the following claims and which will be described in greater detail hereunder. Reference is also being made to Biochemistry 21 , 1982, pages 3298-3303, which discloses the sequence of the natural occuring apoSAA (a precursor of the tissue amyloid protein AA). Reference is furthermore being made to Chemical Abstract 107, 1987, page 308; 107: 37578, Amyloidosis (Proc. Int. Symp. Amyloidosis: Diss. Complex) 4th 1984, pages 407-414, and Chemical Abstract 87, 1977, page 431; 87: 20426t, J. Lab. Clin. Med. 89 (5), 1977, pages 1092-1104. DISCLOSURE OF INVENTION A peptide compound of the type which is specified in the preamble to claim 1 is thus produced in accordance with the invention. The characteristic feature of the peptide compound is that the number of amino-acid residues is at least 6 and in that the said sequence is built up of the following amino-acid residues: Ser-Asp-Ala-Arg-Glu-Asn-Ile-Gln-Arg. The expression "hydrophilicity value" is accepted in the field and is used in accordance with the invention exactly in the manner as described in Proc. Natl. Acad. Sci. USA, vol.78, no. 6, pp. 3824-3828, June 1981. In this literature reference is also found to a table of such values for particular amino-acid residues. The reason why the number of amino-acid residues in the peptide compound in accordance with the invention should be greater than 6, is explained more fully, for example, in the Nature, vol. 299, 14th October 1982, p. 596 for synthetic peptide compounds in general. The peptide compound in accordance with the invention can be prepared synthetically by using in principle conventional methods in the field. Even though it doesn't in general involve any major difficulties to synthesize a peptide compound with a predetermined sequence of amino-acids, it is none the less surprising that a synthetic peptide compound in accordance with the invention can be used for the production of a specific antiserum for AA and/or SAA, bearing in mind that it is particularly difficult when the natural proteins are to serve as an antigen, on the one hand because it is difficult to isolate them wholly free from any contaminating substances, on the other hand because they are difficultly soluble or insoluble in ordinary buffers. Moreover, the peptide conceivable for immunization can be chosen in many differnt manners, since no completely reliable method exists for choosing the "right" peptide. A peptide compound prepared by synthesis in accordance with the invention has been found to function well for precisely the preparation of specific antisera for AA and/or SAA. Said peptide compound is a nonapeptide with the following amino-acid sequence: Ser-Asp-Ala-Arg-Glu-Asn-Ile-Gln-Arg where, in known manner, the codes used have the following meanings Ser = serine Asp = aspartic acid Ala = alanine Arg = arganine Glu = glutamic acid Asn = asparagine Ile = isoleucine Gln = glutamine In accordance with the invention an antiserum is produced also for the detection of AA in a tissue specimen. The antiserum is characterized in that it is procuded by using a petide coumpund of the type described above, i e the nonapeptide of the following amino-acid sequence: Ser-Asp-Ala-Arg-Glu-Asn-Ile-Gln-Arg Furthermore, an antiserum for the determination of the concentration of SAA in blood is produced in accordance with the invention which is characterized in that it is produced by using a peptide compound of the type described above, i e a nonapeptide with the following amino-acid sequence: Ser-Asp-Ala-Arg-Glu-Asn-Ile-Gln-Arg The nonapeptide defined above, coupled to a high molecular carrier was injected in accordance with conventional technique into five rabbits, two times at three weeks' interval. All the rabbits produced antisera which reacted specifically with AA in body tissues as well as with SAA in blood in immunohistochemical techniques and in immunoblood techniques of known type. The production of antiserum continued without loss of titre during six weeks without renewed injections of nonapeptide carrier complex and therefor can be regarded as stable. INDUSTRIAL APPLICABILITY Peptide compounds in accordance with the invention are specially useful for the production of specific antisera for AA and/or SAA. Antisera prepared by using peptide compounds in accordance with the invention are therefore specially useful for the detection of AA in tissue specimens and/or determination of SAA concentration in blood.

Claims: A peptide compound containing amino-acid residues which occur naturally in amyloid A protein and/or serum amyloid A protein, characterized in that the number of amino-acid residues is at least 6 and in that the said sequence is built up of the following amino-acid residues: Ser-Asp-Ala-Arg-Glu-Asn-Ile-Gln-Arg. Use of the peptide compound in accordance with claim 1 for the preparation of antisera. An antiserum for the detection of amyloid A protein in a tissue specimen, characterized in that it is produced using a peptide compound in accordance with claim 1. An antiserum for the determination of the concentration of serum amyloid A protein in a blood sample, characterized in that it is produced using a peptide compound in accordance with claim 1.

Inventors: GRUBB, ANDERS, HANSSON, HELENE, LJUNGGREN, LENNART, Hansson, Heléne
Applicants: GAMBRO LUNDIA AB
Application year: 1986
Main classification: C07K7
IPC codes: A61K39, C07K16, G01N33, C07K14, C07K7