Tips & Tricks

Immunoassays
Poor precision between duplicates (high CV values)
Automated plate washer is not functioning properly and is not washing all wells in the same manner:
Inadequate plate washing:
Problem with the multi-channel pipette during the addition of the conjugate or substrate:
Carry over from high and low samples:
Poor pipetting technique:
Poor standard curve, does not match certificate of analysis
Inadequate plate washing:
Poor pipetting technique:
Inadequate mixing of reagents:
Inadequate shaking of plate:
Wrong volume of reagents added to the well:
Low or inconsistent ambient temperature:
All wells turned bright blue
Automated plate washer is not functioning properly and is not washing all wells in the same manner:
Inadequate plate washing:
Contamination of the substrate with enzyme conjugate:
High background OD readings (Blank or 0 STD values are too high)
Automated plate washer is not functioning properly and is not washing all wells in the same manner:
Inadequate plate washing:
Old or contaminated wash buffer:
Contamination of the substrate with enzyme conjugate:
Wrong conjugate dilution:
Wrong filter used in the plate reader:
Elevated incubation or ambient temperatures:
Unexpected results
Improperly prepared sample:
Omission of a reagent:
Dilution error:
Sample IDs mixed up:
Wrong curve fit used:
No color development or very low OD readings
Wrong filter used in the plate reader:
Inadequate mixing of reagents:
Inadequate shaking of plate:
Reagents contaminated or added in the wrong order:
Assay incubation times not followed correctly:
Improperly prepared sample:
Low ambient temperature:
Control out of range
Reagent stability issue:
Standard curve issue:
Incorrect reconstitution volume used:
Wrong curve fit used:
Flow Cytometry
Reducing non-specific binding
Non-specific binding can be due to several reasons.
Infinicyt™: Flow Cytometry Software
HPLC/LC-MS
No peaks
Double peaks
Contaminating peaks
Variable retention times
Baseline is drifting
Baseline is not smooth
Chemiluminescence
Chemiluminescent Plate Reader Settings
Reader Settings