A Novel Approach to Single Cell RNA-Sequence Analysis Facilitates In Silico Gene Reporting of Human Pluripotent Stem Cell-Derived Retinal Cell Types.

Cell type-specific investigations commonly use gene reporters or single-cell analytical techniques. However, reporter line development is arduous and generally limited to a single gene of interest, while single-cell RNA (scRNA)-sequencing (seq) frequently yields equivocal results that preclude definitive cell identification. To examine gene expression profiles of multiple retinal cell types derived from human pluripotent stem cells (hPSCs), we performed scRNA-seq on optic vesicle (OV)-like structures cultured under cGMP-compatible conditions. However, efforts to apply traditional scRNA-seq analytical methods based on unbiased algorithms were unrevealing. Therefore, we developed a simple, versatile, and universally applicable approach that generates gene expression data akin to those obtained from reporter lines. This method ranks single cells by expression level of a bait gene and searches the transcriptome for genes whose cell-to-cell rank order expression most closely matches that of the bait. Moreover, multiple bait genes can be combined to refine datasets. Using this approach, we provide further evidence for the authenticity of hPSC-derived retinal cell types. Stem Cells 2018;36:313-324.

Surgical sponge blood salvage spinning device design and testing.

One method to clear intraoperative blood from the surgical field is to remove blood with surgical sponges. Currently, absorbed blood cannot be retrieved effectively and is lost. A spinning device was created to salvage red blood cells from the sponges. With this device the sponges are gently washed with saline and the resultant bloody fluid can be delivered to a cell saver to prepare it for autologous blood transfusion. In this article, we demonstrate how a novel sponge extractor can be used to extract blood from sponges. Several tests were conducted with porcine blood to optimise viable blood salvage by varying spin speed, and spin time of the device. At spin speeds greater than 1000 RPM, the blood salvaged from the device was similar to blood volumes obtained by hand wringing sponges. Cell viability testing yielded no significant differences in haemolysis for device trials compared to gently hand wringing. Spin time testing showed no significant differences in the blood salvaged at times greater than one minute. Optimal parameters for the device were determined to be a one-minute spin time at 1500 RPM.