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Troubleshooting Immunoperoxidase Assays |
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Problem: Positive Control is negative or very weak.
Possible Causes: |
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| 1. | Please note that Controls available from Fuller Laboratories have been bottled at "screening dilution". This dilution depends upon the specific assay, but in all cases Positive Controls are intended to be used initially without dilution to demonstrate a "strong" reaction. These Controls give endpoint titers 8-fold beyond the screening dilution (range 1:4-1:16). |
| 2. | If IFA Substrate Slides are purchased as components, the Positive and Negative Controls available from Fuller Laboratories may be used to locate the optimal conjugate titer for use. This optimal titer should give a clear 1+ positive reaction at a Positive Control dilution of 1:8 (see #1 above) and a clearly negative reaction using the undiluted Negative Control. |
Questions / Problems
- Positive Control is negative or very weak.
- Antigen or substrate cells smeared or absent from wells before test.
- Antigen or substrate cells washed off during the procedure.
- Negative Control is positive.
- Reactions are stronger on the edges of the well in comparison to the center.
- What reaction density (bacteria) constitutes a positive reaction?