Counting cells using flow cytometry should pay attention to the problem

Tissue homogenates or cell suspensions are prepared and stained with fluorescent dyes, and the number of cells with different DNA content or multiples can be counted by flow cytometry. For example, in testicular tissue, sperm cells are haploid cells, spermatogonia, Sertoli cells, Leydig cells, and other non-spermatogenic cells are diploid cells, primary spermatocytes, and spermatogonia and non-dividing sperm cells. The spermatogenic cells are tetraploid cells. Therefore, the use of flow cytometry to determine changes in spermatogenic cells DNA DNA ploidy, can indirectly determine the relative proportion of different spermatogenic cell changes in the trend. However, be cautious when using these results to draw conclusions. For example, a decrease in the percentage of haploid cells may indicate a decrease in the number of sperm cells, but the following three conditions may also cause a decrease in the percentage:

One is that the number of haploid cells does not change and the number of other ploidy cells increases;

Second, the number of haploid cells increased, and the number of other ploidy cells also increased and increased more;

Third, the number of haploid cells decreased, and the number of other ploidy cells also decreased, but only a relatively small decrease.

Similarly, the same percentage may indicate that spermatogenesis has not been affected, but the following two conditions may also lead to such results: haploid cells and other ploidy cells increase in proportion; haploid cells and other ploidy The proportion of cells decreases. Therefore, irrespective of the exact identification of cells and the problem of cell comminution during tissue homogenization, flow cytometry estimates the cell number has at least one defect: it generally reflects only the relative number ratio of cells, and cannot determine the measured cells at a certain level. Does the total number of areas of interest really increase, decrease, or change?