An analysis of optimal cell counting range of the LUNA-FX7™
#LOQ #Limit of Quantification #optimal cell counting range #detection range #dynamic range
#wide range of cell concentrations #counting reliability #convenience #performance
HL60 cells of 2.50E+07 cells/mL and the serial dilutions were prepared to be counted in the Brightfield and Fluorescence Cell Counting modes of the LUNA-FX7™. Prior to counting, the cells were stained with the 0.4% Trypan Blue Stain (Cat# T13001) and loaded in the 2-channel LUNA™ Cell Counting Slides (Cat# L12001) for brightfield counting. For fluorescent counting, the Acridine Orange/Propidium Iodide Stain (Cat# F23001) and the 2-channel PhotonSlide™ (Cat# L12005) were used. The conventional hemocytometer count was performed to cross-check the count results of the LUNA-FX7™. For determining the capacity of LOQ ranges of the LUNA-FX7™ Automated Cell Counter, the typical cell size comparable beads (12 µm) were applied in both the Brightfield and the Fluorescence Cell Counting modes using three types of slides, 1-channel, 2-channel, and 8-channel slides*1. The default protocols were adopted for all the counts, and the LOQ ranges were statistically evaluated based on the differences of measured vs. expected counts, combined with co-efficiency of replications. To further verify the higher concentrations, the beads of 7 µm were counted on the Fluorescence Cell Counting mode.
- *1 1-channel: LUNA™ 1-Channel Slides (Cat# L72011), 2-channel: LUNA™ Cell Counting Slides (Cat# L12001) for brightfield counting, PhotonSlide™ (Cat# L12005) for fluorescence counting, 8-channel: LUNA™ 8-Channel Slides (Cat# L72001)
- Figure 1. The comparable counts of HL60 cells: the LUNA-FX7™ vs. a hemocytometer. The comparable cell concentrations (A) and the corresponding table (B) show comparable cell count values of the LUNA-FX7™ and a hemocytometer. The results in both counting modes of the LUNA-FX7™ are not significantly different from that of the standard manual count with a hemocytometer.
- Figure 2. The images of HL60 cell counting in serial dilutions. The brightfield raw images and corresponding fluorescent-tagged images express the counting status from the concentration of 1.56E+06 cells/mL to 2.50E+07 cells/mL.
- Figure 3. The LOQ ranges of slide options. The plots correspond to counting limits of 1-ch, 2-ch, and 8-ch slides in the LUNA-FX7™, and the blue boxes with dashed lines indicate the suggested ranges of LOQ for the slide option and the mode applied (A). The suggested ranges of LOQ of each slide type are defined with numbers in the table (B). Specifically, the 1-ch slide shows the most comprehensive range among the three slide options.
- Table 1. Limit of quantification of slide options on brightfield and fluorescence counting.
- Figure 4. Limit of quantification of small-sized bead counts. In the smaller bead counting (7 µm), the raw images and the corresponding tagged images in the Fluorescence Cell Counting mode display more comprehensive ranges to the highest concentration (5.03E+07 cells/mL).
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