126 | Chapter 4: Instrument Qualification
BioTek Instruments, Inc.
Repeatability Test
1 For each well, calculate the mean and standard deviation of the five
readings.
2 For each mean calculated in step 1, calculate the allowed deviation
using the repeatability specification for a 96-well plate in the Normal
Read Mode, which is ± 1.5% ± 0.005 OD from 0.000 to 2.000 OD @
340 nm (mean * 0.015 + 0.005).
3 For each well, compare the standard deviation calculated in step 1 with
the allowed deviation calculated in step 2. The standard deviation
should be less than the allowed deviation.
For example:
Absorbance readings of 0.802, 0.802, 0.799, 0.798, and 0.801 will result in
a mean of 0.8004 and a standard deviation of 0.0018. The mean
multiplied by 1.5% (0.8004 * 0.015) equals 0.012, and when added to the
0.005 (0.012 + 0.005) equals 0.017, which is the allowed deviation for
well A1. Since the standard deviation for well A1 is less than 0.017, the
well meets the test criteria.
Linearity Test
1 For each of the three dye concentrations, calculate the mean absorbance
for the wells containing that solution (mean of wells A1 to H2, A3 to
H4, and A5 to H6).
2 Perform a regression analysis on the data to determine if there is
adequate linearity.
Example using Microsoft
®
Excel:
In a spreadsheet, enter the three mean values in ascending order and
label the column as the Y values. Enter 0.50, 0.75, and 1.00 and label the
column as the X values.
Select
Tools|Data Analysis|Regression. Identify column Y as the
“Input Y Range” and column X as the “Input X Range” and then click
OK to perform the analysis.
If the Data Analysis command is not available on the Tools menu, you
may need to install the Analysis ToolPak in Microsoft
®
Excel. Consult
Excel’s help system for assistance.
Expected Results:
Since it is somewhat difficult to achieve high pipetting accuracy when
conducting linear dilutions, an R-Square value greater than or equal to
0.990 is considered adequate.
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