Scl-70, Bovine (Scleroderma, DNA Topoisomerase I)

Molecular Biology / MB-Topoisomerases

-20°C/-70°C

Supplied as a liquid in 20mM Tris HCl, 0.60M sodium chloride, 0.1mM DTT, pH 7.4, 0.1mM PMSF, 20% glycerol and 0.1% bromo-nitro-dioxane/methylisothiazolone.

Topoisomerase I changes the topology of DNA via cleavage of single strand only. Suitable for use in DNA conformational analysis and topology as well as DNA repair, drug resistance, cell proliferation and leukemia studies. The double-helical configuration that DNA strands naturally reside in makes them difficult to separate, and yet they must be separated if enzymes are to transcribe the sequences that encode proteins, or if chromosomes are to be replicated. In so-called circular DNA, in which double helical segment is bent around and joined in a circle, the two strands are topologically linked, or knotted. They cannot be separated by any process that does not involve the breaking of strands. Topoisomerases catalyze and guide the unknotting of DNA.

Type I topoisomerases cut only one strand of DNA; type I topoisomerase of E. coli > E. coli (omega protein) relaxes negatively supercoiled DNA and does not act on positively supercoiled DNA. Type II topoisomerases cut both strands of DNA; type II topoisomerase of E. coli (DNA gyrase) increases the degree of negative supercoiling in DNA and requires ATP. It is inhibited by several antibiotics, including nalidixic acid and ovobiocin.

Antibodies generated against the nuclear constituents are known as antinuclear antibodies (ANA). This includes autoantibodies directed against the extractable (soluble in physiological buffers) nuclear antigen or ENA. The most prominent of ANAs/ENAs are autoantibodies which binds to ds-DNA, ss-DNA, histones, ribonucleoproteins (RNP) and the SS-A, SS-B, Sm antigens, Jo-1, and Scl-70. Two antibodies, anti-dsDNA and anti-Sm, appear to occur only in SLE. Others occur in a variety of autoimmune and mixed connective tissue diseases.

Antibody to Scl-70 antigen was originally called Scl-1. Anti-Scl-70 has been shown to react with a cellular antigen known as DNA topoisomerase I, which is responsible for the relaxation of supercoiled DNA. Anti-Scl-70 is found in ~75% patients with the diffuse progressive form of scleroderma. The presence of anti-Scl-70 seems to exclude the presence of anti-centromere antibody, which is a marker for a subset of patients with scleroderma known as CREST (calcinosis, sclerodactyly, and telangiectasia) syndrome.

The frequency of ANA-positives in various rheumatic diseases has been reported for SLE, rheumatoid arthritis (RA), progressive systemic sclerosis (PSS), polymyositis (PM), dermatomyositis (DM), mixed connective tissue diseases, drug-induced SLE, and Sjogren’s syndrome (SS). Most of these studies are based on tedious fluorescent ANA (FANA). Other techniques such as RIA, immunodiffusion, hemagglutination, electrophoresis and immunoblotting are also used to define antibody specificity. Recently, immunological assays (mostly ELISA) that determine the specificity of ANA have been used in studying patients with systemic rheumatic diseases.

Applications:
Suitable for use in ELISA. Other applications not tested.

Recommended Dilution:
ELISA: 0.5-1.0ug/ml coating concentration
Optimal dilution determined by the researcher.

Storage and Stability:
May be stored at 4°C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20°C. Aliquots are stable for 6 months after receipt at -20°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.