Microplates (also called “microtitre,” “microtiter ®,” or “micro titration” plates) provide a rapid way to analyze large numbers of tests. Samples are typically on the order of a few hundred µL, and a large number of plates can be measured in a short period of time. A 96 well plate is commonly used for these analyses, however 12, 24, 48, 384, and 1536 well plates are also available.

A broad variety of analytical measurements can be performed in microplates including absorbance (OD), luminescence (L), fluorescence intensity (F), time-resolved fluorescence (TRF), and fluorescence polarization (FP). In each case, the fundamental measurement is the same as when a standard cuvette is used, but the instrument (called a “reader”) is capable of reading the entire plate is less than a minute and generate a complete report.

While the general design of a reader is identical to that of a cuvette system (e.g. an absorbance reader is similar in nature to a spectrometer), the optics are configured such that 96 samples can be read on a plate with an 8 x 12 format.

A QC Pak™ consists of a plate with a number of compounds to evaluate the performance of the reader, and an Excel template that calculates a variety of parameters such as linearity, crosstalk between wells, and alignment of the plate in the reader. A report is automatically generated which can be printed and/or archived.

This manual describes the use of ABS QC Pak for filter and monochromator-based microplate readers.

The minimum computer requirements for QC Pak are:

Microsoft Windows 2000

Intel Pentium III 500 MHz

128 MB RAM

100 MB Free Hard Disk

Excel 97

 

Figure 1: Raw Data Sheet

The analytical data can be viewed by selecting the tabs on the bottom of the spreadsheet:

The X-Talk and Alignment Tab presents:

• Averaged Data

The averages of the five measure-ments with 405nm, as well as the results of the alignment of the microplate transport. The gray cells indicate the background, which is from empty cells on the plate.

 

Figure 2: Averaged Data

• Microplate Alignment Plot

The Microplate Alignment Plot (Figure 3) presents the ansorbance readings for Column 1 and Column 12 for rows 1 - 8.

The cells in column 1 and column 12 of the plate are the same, and the two lines will coincide if the tray is properly aligned.

Figure 3: Microplate Alignment Plot

• Alignment and X-talk Data & Results

The Alignment and Cross-Talk Results

(Figure 4) describe the statistical analysis of the data and presents the results for the Alignment and Cross Talk tests.

Figure 4: Alignment and Cross-Talk Data and Results

 

• The acceptance criteria are:
• Alignment Column 1 – slope is less than 5 %
• Alignment Column 2 – slope is less than 5%
• Alignment Left to Right – the average 1 & 2 within 20%
• Crosstalk, Best Case – less than 3 x 10 -5
• Crosstalk, Worst Case – less than 1 x 10 -2

The Linearity Tab presents:

Averaged Data

same as in “ X-Talk and Alignment Tab”

Linearity Plot

The Linearity Plot (Figure 5) presents the regression line of the averages for each of the 5 readings at 405 nm of the 7 neutral density filters (column 2 – 8, row 1 – 4 each)

 

Figure 5: Linearity Plot

 

Data Region

The Data Region (Figure 6) presents a summary of the analytical data

Figure 6: Data Region

Results Field

The Results field (Figure 7) indicates the results of the various tests.

Figure 7: Results Field

The acceptance criteria are:

• Linearity Relative to Nominal - R 2 should be greater than 0.900
• Same Well Precision (High Concentration) - should be less than 10%
• Same Well Precision (Low Concentration) - should be less than 0.010
• 405: Reading of 405nm QC filter must be lower 1.0
  Blocking on other QC filters must be higher 4.0
• 450: Reading of 450nm QC filter must be lower 1.0
  Blocking on other QC filters must be higher 4.0
• 492: Reading of 492nm QC filter must be lower 1.0
  Blocking on other QC filters must be higher 4.0
• 620: Reading of 620nm QC filter must be lower 1.0
  Blocking on other QC filters must be higher 4.0