Cross-Talk Test - tecan GENios Operating Manual

7.3.2

Cross-Talk Test

June 2002 Operating Manual for GENios, GENios FL, and GENios Plus No: I 112 904 Rev No: 1.1
Cross talk is a measure for increase of blank values (blank contains pure buffer
and substrate) due to optical leakage from neighboring wells.
•
Greiner 96 well plate, flat bottom, white using black cover plate.
•
A single well (for example: E5) is supplied with 20µl of 0.3 10
units/µl alkaline phosphatase dilution and with 100µl of substrate
•
Fill all other wells with pure buffer and substrate.
1 2 3 4 5
B B B B B
A
B B B B
B B
B
B
B B B
C
B
B B
D N
B
B
B B
E N
B
B B B
N
F
B B
B B
G B
B
B B B B
H
•
Select maximum dynamic range and adjust gain to get a
RLU (Relative Luminescence Units) reading greater than 10
Measurement should be performed 10 minutes after preparation but not later
than 30 minutes.
Calculation method:
Compute MVB (B) from all wells except the nearest neighbor wells (N) and the
well filled with alkaline phosphatase (for example: E5).
Calculate the SD (Standard Deviation) of MVB. Use factor 3 to get higher
statistical reliability.
Calculate the MVN (Mean Value of the Neighbors), using the wells indicated with
the letter N.
If MVN is within the interval MVB +/- 3 SD, cross talk cannot be clearly
distinguished from the background.
If MVN is higher than MVB +3 SD the cross talk CT (Cross Talk) calculates
according to:
CT = (MVN – MVB ) / RLU reading
CT is expected to be ≤ 0.0001
6 7 8 9
10 11
B
B B B B
B
B B
B
B B B
B B
B B B B
B B B B
B B
N B B B B B
B B B
B B B
B B
B B B B
B B B B B B
7. Quality Control
-3
activity
12
B
B
B
B
B
B
B
B
6
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7-5