Anti-Double Stranded DNA (dsDNA) BioAssay™ ELISA Kit, Semi-Quantitave
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The United States Biological Anti-dsDNA ELISA Kit is a quantitative assay intended for the detection of antibodies to dsDNA an†igen in human serum as an aid in diagnosis of Systematic Lupus Erythematosus (SLE). This kit is intended for research use only.
PRINCIPLE OF THE ASSAY:
The dsDNA test is an Enzyme-Linked Immunosorbent Assay to detect antibodies to dsDNA antigen. Purified dsDNA antigen is attached to a solid phase microassay well. Enzyme-Linked Immunosorbent Assays (ELISA) rely on the ability of biological materials (i.e., antigens) to adsorb to plastic surfaces such as polystyrene (solid phase). When antigens bound to the solid phase are brought into contact with serum sample, antigen specific antibody, if present, will bind to the antigen on the solid phase forming antigen-antibody complexes. Excess antibody is removed by washing. This is followed by the addition of goat anti-human IgG and IgM conjugated with horseradish peroxidase which then binds to the antibody-antigen complexes. The excess conjugate is removed by washing, followed by the addition of Chromogen/Substrate, tetramethylbenzidine (TMB). If specific antibody to the antigen is present in the serum sample, a blue color develops. When the enzymatic reaction is stopped with 1N H2SO4, the contents of the wells turn yellow. The color, which is indicative of the concentration of antibody in the serum, can be read on a suitable spectrophotometer or ELISA microwell plate reader.
D3878-14A1:. Microtiter Plate, 12x8 strips. dsDNA antigen-coated wells, sealed in a resealable foil pouch with desiccant.
D3878-14A2: Serum Diluent Type II, 1x30ml. Ready to use. Contains proclin (0.1%) as a preservative.
D3878-14A3: Positive Control, 1x0.4ml. Human serum or defibrinated plasma. Sodium azide (< 0.1%) and pen/strep (0.01%) added as preservatives, with established range printed on vial label. The Positive Control is utilized to control the positive range of the assay. (96T: one vial, 0.4 mL)
D3878-14A4: Calibrator, 1x0.4ml. Human serum or defibrinated plasma. Sodium azide (< 0.1%) and pen/strep (0.01%) added as preservatives, with kit specific factor. The Calibrator is used to calibrate the assay to account for day-to- day fluctuations in temperature and other testing conditions.
D3878-14A5: Negative Control, 1x0.4ml. Human serum or defibrinated plasma. Sodium azide (< 0.1%) and pen/strep (0.01%) added as preservatives, with established range printed on vial label. The Negative Control is utilized to control the negative range of the assay.)
D3878-14A6: Horseradish-peroxidase (HRP) Conjugate, 1x16ml. Ready to use. Goat anti-human IgG and IgM, containing proclin (0.1%) and gentamicin as preservatives.
D3878-14A7: Wash Buffer Type I (20X concentrate), 1x50ml. Dilute 1 part concentrate + 19 parts deionized or distilled water. Contains TBS, Tween-20 and proclin (0.1%) as a preservative.
D3878-14A8: Chromogen/Substrate Solution Type I, 1x15ml. Tetramethylbenzidine (TMB), ready to use. The reagent should remain closed when not in use. If allowed to evaporate, a precipitate may form in the reagent wells.
D3878-14A9: Stop Solution, 1x15ml. Ready to use, contains a 1N H2SO4 solution.
MATERIALS REQUIRED BUT NOT PROVIDED:
1. Wash bottle, automated or semi-automated microwell plate washing system.
2. Micropipettes, including multichannel, capable of accurately delivering 10-200ul volume (less than 3%CV).
3. One liter graduated cylinder.
4. Paper towels.
5. Test tube for serum dilution.
6. Reagent reservoirs for multichannel pipettes.
7. Pipette tips.
8. Distilled or deionized water (dH20), CAP (College of American Pathology) Type 1 or equivalent.
9. Timer capable of measuring to an accuracy of +/- 1 second (0 - 60 minutes).
10. Disposal basins and 0.5% sodium hypochlorite (50 mL bleach in 950 mL
11. Single or dual wavelength microplate reader with 450 nm filter. If dual wavelength is used, set the reference filter to 600-650 nm. Read the Operator's Manual or contact the instrument manufacturer to establish linearity performance specifications of the reader.
Note: Use only clean, dry glassware.
STORAGE AND STABILITY:
Store all components at 4°C. Stable for 6 months. For maximum recovery of product, centrifuge the original vial prior to removing the cap.
Important Note: This product as supplied is intended for research use only, not for use in human, therapeutic or diagnostic applications without the expressed written authorization of United States Biological.