A reaction mix was prepared manually using 5 parts of 2x Rotor-Gene SYBR Green PCR Master Mix and 3 parts of RNase free water (ready-made reaction mix). An experiment was then defined in the QIAgility Setup Manager for the Rotor-Gene SYBR Green PCR Kit with a total reaction volume of 25 µl and a sample input volume of 5 µl, resulting in the transfer of 20 µl of the ready-made reaction mix. The transfers were made into pre-weighed 200 µl single tubes (n=36 per instrument). The tubes were weighed again after the transfer and the weight difference was considered to calculate the transferred volume. For each instrument, the pipetting precision was determined as CV (%). The data shows the mean of the CV values (bars; in %) ± S.D. (error bars) of each individual instrument. The pipetting precision or CV, respectively, was nearly 3 % for all instruments tested.
The indicated CV values (%) demonstrate that the QIAgility instrument has high performance rate in terms of precision and repeatability when pipetting ready-made reaction mix as shown on all 4 instruments.
Human gDNA with a stock concentration of 1.5 µg/µl was diluted with the QIAgility six times in the ratio of 1:2 per dilution step using RNase free water as diluent. The dilution series was prepared in replicates of 3. The nucleic acid concentration of the dilutions was determined using the QIAxpert UV/VIS spectrophotometer (DNA QIAamp Application). The mean measured quantifications were plotted against the mean theoretical nucleic acid concentrations assuming a perfect 1:2 dilution. The resulting plot demonstrates the exceptional linearity of QIAgility dilution with a high regression coefficient of >0.999.
Quantifications of serial dilutions of gDNA prepared using the QIAgility are in high concordance with actual concentrations.
A QuantiNova SYBR Green assay setup was performed using the QIAgility operated with its Setup Manager Software and this was compared to a manual setup done in parallel. A manually prepared 1:10 dilution series from a stock of human gDNA with a concentration of 60 ng/µl was used in a ß-actin assay. Each dilution was tested in quadruplicates (n=4). The manual and QIAgility-prepared assay points were amplified in the same Rotor-Gene Q run. The data is represented as a box plot of the CT value calculated using the Q-Rex Software for each template concentration and setup employed.
A highly similar PCR performance is observed between manual and automated assay setup over a series of sample concentrations.
The Artel MVS system was employed to determine the pipetting precision of the QIAgility instrument operated by the QIAgility Setup Manager Software. The QIAgility dispensed the Artel solutions (selected according to the Artel MVS User Manual) into the Artel buffer. Three runs were performed on three QIAgility instruments that were used to pipet an entire volume range (1, 2, 5, 10, 20, 50, 100, and 200 µl), in replicates of 12. For each run and volume (n=12) the coefficient of variation (CV) was calculated to determine the pipetting precision of the QIAgility. The data shows the mean of the CV values (bars; in %) ± standard deviation (S.D.) of the CV values as error bars. The red lines depict the claimed specifications.
The indicated CV values (%) demonstrate that the QIAgility instrument performs within its claimed specifications and even better.