AUTHOR=Yaigoub Hasnaa , Tirichen Hasna , Xin Xiaohong , Shi Shuhong , Wu Changxin , Li Rongshan , Li Yafeng 

TITLE=Isolation of Viable Single Cells With High Yield and Purity Using a Small Amount of Human Kidney Tissue Biopsy

JOURNAL=Frontiers in Cell and Developmental Biology

VOLUME=Volume 10 - 2022

YEAR=2022

URL=https://www.frontiersin.org/journals/cell-and-developmental-biology/articles/10.3389/fcell.2022.822275

DOI=10.3389/fcell.2022.822275

ISSN=2296-634X

ABSTRACT=Objective: Establishment of an efficient method of preparing human kidney single-cell suspension, using a very small amount of tissue puncture. Methods: Samples of human kidney tissue puncture were cut into pieces, and then 80 μL of the digestive enzyme were added to each punctured tissue to induce enzymatic digestion. The enzyme combination is composed of collagenases, DNase and hyaluronidase and the sample was incubated 20 minutes at 37 ℃. The obtained cell suspension was filtered through a 70 μm cell strainer, centrifuged at 300g for 5 minutes and the supernatant was removed, then the pellet was resuspended in 3 mL of DMEM (Dulbecco's Modified Eagle's Medium). Cell suspension was sorted and purified by flow sorting to remove dead cells and obtain a cell suspension with higher viability rate. Results: We found that (1) diverse single cells of human kidney can be obtained by the digestive enzyme, as observed under the light microscope, with different sizes, normal cell morphology and good dispersion. (2) (2-3)×106 single cells can be extracted from one fresh punctured kidney tissue of about 10mg, with a cell viability rate of more than 80%. Conclusion: In this work we generated a comprehensive and high-resolution single-cell method, which is simple and efficient for preparing single-cell suspension from a minimal amount of human kidney tissue. This method can facilitate the study of renal cell biology and the pathogenesis of kidney diseases.