MICROPLATE READERS
Fig. 1. Detection of diff erent ratios of WT-HeLas and GFP+ VantaStar, using fl uorescence matrix scans and the bottom optic
well with diff erent ratios from 0 (only WT-HeLas) to 100% (only GFP+ mcherry+
/
-HeLas) were seeded in 96-well plates. After cellular attachment, cell nuclei of all cells were additionally stained with the fl uorescent dye Hoechst33342.
/mcherry+
-HeLas with the
As displayed in fi gure 1, GFP, mcherry and Hoechst fl uorescence signal intensity were measured on the VantaStar using a 15 x 15 matrix scan with a diameter of 6 mm. T e use of the bottom optic allows to reduce background derived from auto-
fl uorescent medium components with one click. T e included scan options thereby help to reduce data variability derived from heterogeneous samples like adherent cell cultures. T e VantaStar’s LVF Monochromator provides full fl exibility together with high sensitivity. For the measurement of GFP, mcherry and Hoechst33342 fl uorescence intensity, the fl uorophore pre-sets, available with the LVF Monochromator, were used.
As presented in Fig. 2, exemplary for GFP fl uorescence detection, a linear relationship between the percentage of GFP+
/mcherry+ HeLas (= transfection
effi ciency) and the measured signal for GFP fl uorescence is observed with high accuracy (R² = 0,9997) and precision (%CV = 10.5). As visible from the consistent fl uorescence intensities for Hoechst33342, the readout of stained cell nuclei reliably correlates with the total cell counts and is well suited to normalise the GFP+
/mcherry+ signal.
With the Enhanced Dynamic Range (EDR) feature, the gain settings required for the best-possible fl uorescent detection do not have to be determined manually in advance. With EDR each well is automatically read with the ideal gain and the measurement values are set in relation to each other without preceding manual steps. As a multi-mode reader, the VantaStar is also able to assess transfection assays based on luminescence or absorbance. T e VantaStar is an advanced detection platform to monitor transfection effi ciency bypassing the need for the more time- consuming generation and evaluation of microscopic images. As a multi-mode microplate reader, it allows to precisely assess transfection assays in various detection modes.
Fig. 2. Linear relationship of percentage of GFP+ /mcherry+ -HeLas
(=transfection effi ciency) and obtained GFP signal with matrix scan. Error bars refer to 8 replicates. Matrix scan example shown for one well each (round area below the bars)
Ann-Cathrin Volz is with BMG Labtech
www.bmglabtech.com
www.scientistlive.com 21
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