La （SSB） antigen

Identity:La ribonucleoprotein, SSB antigen, Sjögren syndrome antigen B.Source Material:Bovine thymus (New Zealand origin).Clinical Indications:Autoantibodies present in Sjögren’s syndrome, systemic lu

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La （SSB） ANTIGENAROTEC_La-SSB_Product_Info.pdf Version/Date: B/04.05.20ATL01-02 La （SSB） antigen 0.20 mgATL01-05 La （SSB） antigen 0.50 mgATL01-10 1.0 mg_________________________________________________________________________________Description of the ProductPurified from bovine thymus. After coating onto ELISA platesthe product will bind autoantibodies to La （SSB）.Purity: The La autoantigen （45-50 kDa） is more than 90%pure, as assessed by SDS polyacrylamide gel electrophoresis.Concentration: 0.1-1.0 mg protein/ml.Storage: The product is stabilised with 20% glycerol and 0.1%Micr-O-protectTM. Store at -20 oC or below （long term） or at+4oC （short term）. Avoid repeated freezing and thawing. Mixthoroughly before use.Clinical and Biochemical DataSjögren's syndrome （SS） is a common systemic autoimmuneinflammatory disorder characterised by lymphocyte-mediateddestruction of exocrine glands leading to diminished or absentglandular secretion1-4. SS may present as a primary diseaseor in association with other systemic autoimmune diseases（referred to as secondary SS）. Autoantibodies to the La （SSB）antigen can be detected in the sera of up to 87% of patientswith primary or secondary SS5,6. The presence of anti-La（SSB） autoantibodies usually coincides with the presence ofanti-Ro （SSA） autoantibodies7, however the fact that anti-Roautoantibodies are far more common in other rheumatologicalconditions such as systemic lupus erythematosis （SLE） andmixed connective tissue disease （MCTD） suggests that anti-Lais more specific for primary and secondary SS than anti-Ro8,9.Anti-La autoantibodies have also been reported to be presentin other clinical conditions, most notably in the sera of mothersof infants with neonatal lupus syndrome10, but also in 10 to15% of SLE patients11,12. binds to the oligo（U） 3' termini of nascentRNA polymerase III transcripts and facilitates transcriptionaltermination and reinitiation by this enzyme13,-17. It has alsobeen reported to function as an ATP-dependent helicase ableto melt RNA-DNA hybrids18. La （SSB） may be involved inother processes as well such as maturation and/or nuclearexport of RNA polymerase III products and some aspects oftranslation19,20. La （SSB） is a highly phosphorylated proteinwhich migrates at about 50 kDa in SDS-polyacrylamide gelelectrophoresis21. Phosphorylated residues are present at thecarboxy-terminal part of the protein22. At least 8 isoelectricforms （pI range 6 to 7） have been identified23.The amino acid sequences of both human and bovine La（SSB） antigen have been determined by cDNA cloning andsequencing19,28. Comparison of the two sequences shows 22largely conservative amino acid substitutions with a total of95% identity. Three regions of the La molecule （amino acids1-107, 111-242 and 346-408） are thought to contain the majorepitopes reactive with human anti-La sera19,24. The broadcross-reactivity of patient sera with La （SSB） from diversemammalian species indicates the presence of conservedepitopes25. The use of bovine for thedetection of human anti-La （SSB） antibodies has beendescribed by several authors25-27.MethodologyThe following is an ELISA procedure which can be used todetect anti-La （SSB） autoantibodies in human serum using theATL01 purified autoantigen:1. Dilute the purified antigen to 0.5-1.0 µg/ml in PBS （10 mMpotassium phosphate, pH 7.4, 0.15 M NaCl）.2. Coat ELISA plates with 100 µl of diluted antigen per well.Cover and incubate 24 hours at +4oC.3. Empty the plates and remove excess liquid by tapping on apaper towel.4. Block excess protein binding sites by adding 200 µl PBScontaining 1% BSA per well. Cover and incubate at +4oCovernight.5. Empty plates and apply 100 µl of serum samples diluted1:100 in PBS / 1% BSA / 1% casein / 0.1% Tween20.Incubate at room temperature for 1 hour.6. Empty plates and add 200 µl PBS / 0.1% Tween20 perwell. Incubate 5 minutes then empty plates. Repeat this steptwice.7. Apply 100 µl anti-human IgG-enzyme conjugate（horseradish peroxidase or alkaline phosphatase） diluted inPBS / 1% BSA / 1% casein / 0.1% Tween20 per well andincubate for 1 hour.8. Repeat step 6.9. Add enzyme substrate and stop the reaction whenappropriate.10. Read absorbance in an ELISA spectrophotometer.References1. Molina, R. et al. （1986） Am. J. Med. 80, 232. Bloch, K.J. et al. （1965） Medicine 44, 1873. Fox, R.I. et al. （1986） Arthritis Rheum. 29, 5774. Aziz, K.E. et al. （1992） Aust. NZ J. Med. 22, 6715. Manoussakis, M.N. et al. （1986） Scan. J. Rheumatol. 61, 896. Harley, J.B. et al. （1986） Arthritis Rheum. 29, 1967. Craft, J.E. & Hardin, J.A. （1987） J. Rheumatol. 14 S13, 1068. St. Clair, E.W. （1992） Rheum. Dis. Clin. N. America 18, 3599. Harley, J.B. （1989） J. Autoimmun. 2, 38310. Buyon, J.P. et al. （1989） J. Clin. Invest. 84, 62711. Reichlin, M. （1986） J. Clin. Immunol. 6, 33912. Wiascewk, C.A. & Reichlin, M. （1982） J. Clin. Invest. 69, 83513. Stefano, J.E. （1984） Cell 36, 14514. Gottlieb, E. & Steitz, J.A. （1989） EMBO J. 8, 84115. Gottlieb, E. & Steitz, J.A. （1989） EMBO J. 8, 85116. Maraia, R.J. et al. （1994） Mol. Cell Biol. 14, 214717. Maraia, R.J. （1996） Proc. Natl. Acad. Sci. USA 93, 338318. Bachmann, M. et al. （1990） Cell 60, 8519. Chambers, J.C. et al. （1988） J. Biol. Chem. 263, 1804320. Bachmann, M. et al. （1989） Mol. Cell Biochem. 85, 10321. Pruijn, G.J.M. （1994） Man. Biol. Markers Dis. （Kluwer Acad.Publ.） B4.2/1-1422. Pfeifle, J. et al. （1987） Biochim. Biophys. Acta 928, 21723. Francoeur, A.M. et al. （1985） mol. Cell Biol. 5, 58624. McNeilage, L.J. （1990） J. Immunol. 145, 382925. Chan, E.K.L. & Tan, E.M. （1987） J. Exp. Med. 166, 162726. Zhang, W. & Reichlin, M. （1996） Arthritis Rheum. 39, 52227. Chan, E.K.L. et al. （1986） J. Immunol. 136, 374428. Chan, E.K.L. et al. （1989） Nucleic Acids Res. 17, 2233Micr-O-protect is from Roche Diagnostics GmbH （Mannheim,Germany）.Tween20 is a registered trademark of ICI Americas Inc.NOTE: No patented technology has been used by AroTecduring the preparation of this product.