Journal: Genes

Article Title: Recapture Lysosomal Enzyme Deficiency via Targeted Gene Disruption in the Human Near-Haploid Cell Line HAP1

doi: 10.3390/genes12071076

Figure Lengend Snippet: Loss of GBA and the associated changes in the GBA-KO line. ( A ) Live cell images were recorded on the cultured HAP1 and GBA-KO lines. ( B ) The same numbers of cells of either wild-type HAP1 or GBA-KO line were seeded in a 6-well plate (Day 0). The cells were cultured in a complete medium and counted for 3 consecutive days. The cell counts were mean ± standard deviation ( n = 3). ( C ) Cells that were revived and cultured for at least two passages were stained and imaged by a confocal microscope in the wild-type and GBA-KO lines. The lysosome staining images are overlays of fluorescent and transmitted light images using a confocal microscope (magnification of 400×). Lysosomes were red granules in the cytosolic compartment of both cultured HAP1 lines. ( D ) Flow cytometry analysis showed that the red fluorescent intensity was elevated to 755.85 in the GBA-KO line, as compared to 634.02 in the wild-type HAP1. The background red florescence was 351.83 in the unstained HAP1 cells.

Article Snippet: Cells stained with the fluorescent lysosome staining solution were analyzed using an Accuri C6 Plus Flow Cytometer (BD Biosciences, San Jose, CA, USA), as reported.

Techniques: Cell Culture, Standard Deviation, Staining, Microscopy, Flow Cytometry