Anti Scl70 In Serum

Introduction

Scl-70 antigen is purified from calf thymus, bound to microwells and stabilized for extended shelf life. Diluted patient sera are placed in the microwells & incubated. If anti-Scl-70 antibodies are present, they will bind to the antigen in the microwell. After washing the microwells to remove residual sample, a second incubation with anti-human IgG conjugated to alkaline phosphatase is carried out. Conjugate will bind immunologically to the anti- scl-70 IgG of the antigen- antibody complex, forming a 'sandwich' consisting of:
Conjugate (Enzyme-labeled Anti-human IgG)
Human anti-Scl-70 ( IgG)
Well Coated with Scl-70 antigen
Unbound conjugate is removed in the subsequent washing step. Enzyme substrate is then added to the microwell & if bound conjugate is present, the colorless substrate (p-nitrophenyl phosphate) will be hydrolyzed to form a yellow end product ( p-nitrophenol). The intensity of the color is measured photometrically at 405 nm & is proportional to the concentration of anti-Scl-70 present in the patients sample.

Systemic sclerosis ( scleroderma) is a chronic disease of unknown etiology which affects connective tissue & small arteries. It is characterized by fibrotic & degenerative changes in the skin, blood vessels, & internal organs. Two major subtypes of scleroderma have been recognized. Diffuse scleroderma is a rapidly progressive form of the disease characterized by widespread changes in the skin, & involving the distal & proximal extremities, the trunk, and the viscera. The second subtype is limited scleroderma, also known as CREST syndrome. In this milder form of the disease, skin involvement is limited to the fingers, hands, or face, and involvement of internal organs is often delayed for several decades or longer.
Two autoantibodies have been identified which are present almost exclusively in scleroderma. The first of these, anti-Scl-70 antibody is found in 20-50% of patients with diffuse scleroderma. These antibodies are directed against a 70kDa nuclear protein which has been identified as topoisomerase 1. The second autoantibody, anti-centromere is strongly associated with limited scleroderma, or CREST syndrome. This antibody is directed against the chromosomal kinetochore. The detection of either of these autoantibodies permits a high degree of certainty in the diagnosis of scleroderma, & because they rarely occur in the same patient, they are useful in determining the scleroderma subtype, diffuse or limited (CREST).

Serum is separated from the clot & refrigerated, 2-8 degrees C for short term storage or stored frozen, -20 degrees C, for long term storage. Avoid freeze-thaw cycles. CAUTION: Serum samples should not be heat inactivated, as this may cause false positive results.
No special patient preparation required.

Henry, John Bernard, Clinical Diagnosis amd Management by Laboratory Methods, W. B. Saunders Co., Philadelphia, 1991. pp 891-892.
The Physicians Guide to ENA Testing, Diamedix Corporation, Miami, 1991. pp 1-8.
Summary of Procedure. DiaMedix Corporation, Miami, Oct. 1991. pp 1-8.
Poly-ENA Extractable Nuclear Antigen Assay for Detection of Antibodies to RNP, Sm, and/or SSA (RO), & SSB (LA). Zeus Scientific, Inc., Raritan, New Jersey, 1987. pp 1-5.
Medical Knowledge Self Assessment Program (MKSAP) 14, American College of Physicians, Philadelphia 2006