Description

The Diagnostic Automation ANA ELISA kit is designed to detect IgG class antibodies to a variety of common nuclear antigens in human sera. Wells of plastic microwell strips are sensitized by passive absorption with antigen. The test procedure involves three incubation steps: (1) the properly diluted test sera are incubated in antigen coated microwells. Any antigen specific antibody in the sample will bind to the immobilized antigen. Afterwards the plate is washed to remove unbound antibody and other serum components; (2) the peroxidase conjugated goat anti-human IgG (γ chain specific) is added to the wells and the plate is incubated. The conjugate will react with the antibody immobilized on the solid phase in step 1. The wells are washed to remove unreacted conjugate; (3) microwells containing immobilized peroxidase conjugate are incubated with peroxidase substrate solution. The hydrolysis of the substrate by peroxidase produces a color change. After a period of time the reaction is stopped and the color intensity of the solution is measured photometrically. The color intensity of the solution depends upon the antibody concentration in the original test sample.

Quality control must be observed in using the ANA Screen. Each time the assay is run the Calibrator must be run in triplicate. A reagent blank, negative control and positive control must also be included in each assay. Calculate the mean of the three Calibrator wells. If any of the three values differ by more than 15% from the mean, discard that value and calculate the mean using the remaining two wells. The mean OD value for the Calibrator and the OD values for the Positive and Negative Controls should fall within the following ranges:

1. Negative Control < or = 0.250

2. Calibrator > or = 0.300

3. Positive Control > or = 0.500


Where:


a. The OD of the Negative Control divided by the mean OD of the Calibrator should be < or = 0.9.

b. The OD of the Positive Control divided by the mean OD of the Calibrator should be > or = 1.25.

c. If the above conditions are not met the test should be considered invalid and should be repeated.

The Positive Control and Negative Control are intended to monitor for substantial reagent failure and will not ensure precision at the assay cut-off. Additional controls may be tested according to guidelines or requirements of local, state, and/or federal regulations or accrediting organizations.

There are certain limitations in the ANA ELISA Kit: (1) this test is a diagnostic aid and by itself is not diagnostic. Test results should be interpreted in conjunction with the clinical evaluation and the results of other diagnostic procedures; (2) Positive ANA may be found in apparently healthy people. It is therefore imperative that the results be interpreted in light of the patientâ??s clinical picture by medical authority; (3) SLE patients undergoing steroid therapy may have negative test results; (4) Many commonly prescribed drugs may induce ANA; (5) The DAI ANA Screen ELISA Kit will not identify the specific type of ANA present in a positive specimen. Positive specimens should be tested for individual autoantibodies using more specific reflex tests such as the DAI ENA Profile-6 ELISA Kit, in combination with the DAI dsDNA ELISA Kit. Alternatively, specific autoantibodies may be detected using a variety of methods including immunodiffusion, western blot or multiplexed fluorescent bead based assays such as the AtheNA Multi-Lyte ANA ELISA Kit.