Question: How can I remove red blood cells from my sample?
Answer: Red blood cells can be removed from a sample during sample preparation and data analysis.
Removal of RBCs during Sample Preparation:
If using a Ficoll gradient to isolate PBMCs(i.e., Sepmate, CPT, Ficoll Paque reagent), most RBCs will remain in the lowest fraction of the gradient. Any remaining RBCs can be removed using an RBC lysis buffer. Instructions for RBC lysis can be found in the Isolation of Leukocytes, Bone Marrow, and Peripheral Blood Mononuclear Cells for Single Cell RNA Sequencing Demonstrated Protocol.
If no Ficoll gradient is being used (i.e., looking to retain leukocytes, working with tissue tumor sample), RBC lysis is suggested as these cells may make up a large portion of the total cell number. For whole blood samples, instructions for RBC lysis are available in Isolation of Leukocytes, Bone Marrow, and Peripheral Blood Mononuclear Cells for Single Cell RNA Sequencing Demonstrated Protocol. For RBC lysis in tumor samples, instructions can be found in the Tumor Dissociation for Single Cell RNA Sequencing https://support.10xgenomics.com/single-cell-gene-expression/index/doc/demonstrated-protocol-tumor-dissociation-for-single-cell-rna-sequencing.
Removing RBCs and platelets during data analysis:
RBCs and platelets can be removed during data analysis by using common RBC and platelet gene expression markers. Use Loupe Cell Browser to identify and exclude clusters expressing high levels of RBC markers (HBB, HBA1) and platelet markers (PPBP).
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