If the above problem is identified, it is assumed to arise from a contamination with old PCR products of the PCR-mix or from reagents used to prepare this mix ( ). It is not assumed that there are problems in any of the steps in the detection of the solid phase product by hybridization. One of the two following factors may be the cause to the problem:

• General problems with preparation of PCR mix
• Problems with contamination of old PCR products in preparation of PCR mix

In the detection step after PCR. The following factors can give rise to high background signals in this step:

• Concentration and temperature of the washing buffer used after hybridization are not optimal for the detection probe
• In the wash after hybridization normal Tris wash buffer is used
• Incorrect incubation buffer (e.g. PBS or BSA in buffer) when adding streptavidin-enzyme conjugate
• Incorrect incubation conditions during substrate reaction
• The substrate reaction is not terminated before measuring. Conversion of substrate may continue during reading and yield different results from one end of the MicroWell^® plate to the other as well as