Test Instruction – Pretest 1.

1.1 Homogeneity of the optical module

Aim:
With the use of a special dye solution it is possible to test the homogeneity of the optical
module. Therefore the dye solution has to be tested twice.

Test fluid:
Distilled water
Cibacron Dye solution
Microtiterplate, uncoated

Method:
A ready to use Cibacron Blue solution can be purchased from ORGENTEC (Cibacron Dye
solution, #X-11). The optical density (OD) of this solution will be approximately 2.2 when
tested with the Alegria®. Pipette manually 100 µl of this dye solution in the wells 3 and 4 of a
microplate. 29 test strips have to prepared in this way. The optical density of well 3 and 4 has
to be tested
a) with Alegria® (via Service Software) and
b) with a microtiter plate photometer.

a) Measuring with Alegria®

The prepared test strips containing dye solution have to be placed in the SysTrays.

Attention: Position 5 (SysTray A) requests an empty test strip. This one is the reference for
the optical density.

The handling of Alegria® takes place with the service software „Alegria Developer“, Window
„Service“ (figure 1). The latest software version for the instrument and service must be used .
The buttons „Open/ Close Lid“ (open and shut the door) and „Rotate Carousel“ (rotates the
carousel to the different SysTray positions) are required for loading of the instrument.

To test the OD, please enter:

- position of the test strip: enter in „Strip position“
- activate the button „Run OD“: Start of the OD determination. Some seconds later you
will receive the detected values on the left side of this button in the box for „well 3“
and „well 4“

The input for the measurement of the OD can only be done separately for every test strip.
The test values have to be written down before the next inputs (look at „calculation and
documentation“). Please have in mind that the OD value of one sample falls low continuously
if measured several times.

After all samples are tested, the SysTrays can be taken out of the instrument (Buttons
„Open/ Close Lid“ and „Rotate Carousel“, look above) and the OD measurement can take
place with the Tecan instrument.

version 2.0, date 03.03.2005 1

Test Instruction – Pretest 1.1
Fig. 1: User interface of the software „Alegria Developer“, Window: Service

a) Measuring with a microtiter plate photometer

Microtiter strips measured with the Alegria® instrument have to be placed into a microtiter
plate frame. Measure the OD with the microtiter plate photometer at a wavelength of 650 nm
following the manufacturer’s instructions.

Calibration and documentation:

The Error-Log data has to be stored. The button „Load Error.Log“ of Window „Service“ is
necessary and saves the data in a specific Run folder.

The measured OD values of Alegria and the microtiter plate photometer have to be
transferred to the “Pretest 1.1_analysis.xls” file. Every yellow coloured box has to be filled
out. Succeeding this step you will get an automatic calculation in the orange coloured boxes.
The calculation contains Index values, OD- means, standard deviation as well as the
coefficients of variation (CV).

Evaluation and release :

Criteria of acceptance are coefficients of variation (CVs) < 2,5 %.

If the CV is > 2,5 %, the technician decides about the further procedure.

version 2.0, date 03.03.2005 2