High backgrounds

Washing

Insufficient washing.

Plate washer should be checked and served periodically. When washing manually use a squirt bottle, multi channel- or multipipette, see Manual Wash Instructions.

Contamination of Calibrator 0, Assay Buffer or Conjugate Buffer

Contamination of Calibrator 0, Assay buffer or Enzyme Conjugate Buffer with Calibrators, Controls or samples.

Carefully rinse wells that are still blue after washing. This is done with wash solution using a squirt bottle or pipette.

Contamination of Calibrator 0, Assay Buffer or Conjugate Buffer

Contamination of Calibrator 0, Assay buffer or Enzyme Conjugate Buffer with Calibrators, Controls, purified antigen such as “free” insulin or samples.

Always use clean pipette tips, beakers etc. in order to avoid contamination.

Contamination of Substrate TMB

Substrate TMB might be contaminated with enzyme conjugate.

Containers and pipette tips used for enzyme conjugate solution should be kept separate from the Substrate TMB.

Substrate TMB

Substrate TMB has been exposed to light.

Substrate TMB is light sensitive. Uncovered Substrate will cause increased background if left out (for only a couple of minutes).

Dried solution on the bottom of the plate

Dried washing solution under the wells/plate (on the outside).

Dampen a tissue with alcohol and carefully wipe off the bottom of the plate. Re-measure immediately.