Unique label-free cell analysis technology for imaging analysis without the need for fluorescent dye-labeled cells

Unique label-free cell analysis technology eliminates the need for fluorescent dyes

Label cells for imaging analysis

Introduction

Cell imaging-based analytical techniques generally require labeling with fluorescent dyes, and some fluorescent labels may be toxic to living cells or may only be used for stained cells. Unlabeled cell analysis technology allows researchers to calculate cell numbers and cell confluence without the need for time-consuming and labor-intensive staining procedures or the effects of dyes on normal cell viability, and to obtain quantified cell proliferation and health in the first place. The data. The SpectraMax. i3 multi-function microplate assay platform with MiniMax. 300 cell imaging module features patented StainFree. label-free cell analysis technology. This technique can be applied to cell proliferation, cytotoxicity or other assays without the need for DAPI-labeled nuclei. DAPI acts as a living cell dye to bind DNA, but prolonged labeling can be toxic to cells.

This article compares the difference between cell counting using labeled-free cell analysis techniques and counting with conventional fluorescent dye-labeled nuclei and whole cells. Similarly demonstrate how alternative fluorochrome-labeled by label-free cellular analysis techniques to obtain IC ₅₀ curve of compound treated cells.

Advantage

1. Calculate cell number and cell confluence without fluorescent dye labeling

2. Monitor the cells without affecting their normal growth

3. SoftMax Pro software's intuitive user interface makes imaging setup easier and faster

Cell-counting without label cell analysis

CHO cells were plated in 384-well plates at a density of 8000/well to 250/well and cultured overnight. The next day, imaging was performed using the transmitted light (TL) channel of the SpectraMax MiniMax cell imaging system. To better compare the effects of both unlabeled cell counts and fluorescently labeled cell counts, the same cells were fixed and stained with green whole cell dye or red nuclear dye, and then imaged using the corresponding fluorescent channels (541 nm and 713 nm). .

The label-free analysis technique uses the number of cells calculated after transmission of the light channel. In the SoftMax Pro software, an analysis module for many different types of cells has been pre-set, and the results can be obtained by simply reading the button. The operator can also use the instrument's self-learning function, a logic algorithm of the software, to teach the software to identify cells (Figure 1). Counting individual cells or analyzing cell confluence provides a quick way to monitor cell growth anytime, anywhere without staining it. Users can also choose custom settings according to their needs. The temperature control function of the instrument can maintain the cell imaging process at 37 °C, which can avoid the effect of cell imaging on cell viability under long-term room temperature environment.

After data analysis by SoftMax Pro software, we found that the results of red and green fluorescent channel cell counting were highly consistent with the number of cells obtained by the unlabeled method. Pre-defined nuclear count and whole-cell count analysis modules in the software speed up the analysis. Figure 2 shows that the cell number curves calculated by the two methods almost coincide.

Cytotoxicity assay using label-free cell analysis

HeLa cells were plated at a density of 5000/well in 384-well plates and grown overnight. The next day, several compounds that induced cell death were treated for 72 hours and analyzed using a SpectraMax MiniMax cell imaging system. The number of viable cells after treatment was calculated by label-free analysis, and its IC ₅₀ curve was plotted using Soft-Max Pro software. The results show that the reaction of the cells to the compound can be accurately detected without the destructive staining and decolorization steps (Fig. 3).

to sum up

Label-free analysis technology as a new method for cell counting and monitoring of cell growth status can greatly reduce test time and expensive dyes. No need to fix cells and staining means that living cells in growth can be detected and detected at any time without disturbing the normal growth after the cells, facilitating subsequent analysis and detection.

SoftMax Pro software has a simple imaging analysis interface, which provides a user-friendly preset analysis module for various common cell types. The same researcher can also customize the settings according to his own needs.