We explored a technique called stepwise enzymatic digestion (StE) method for preparing a single-cell suspension of rat whole kidney tissue which contained three main steps. The first step is to cut the kidney into a homogenate. The second step is the digestion of renal tubules using Multi ...
By using this method, the difficulty of the procedure was reduced, fewer cells were lost during the preparation treatments, and the maximal activity of single cells was maintained. Following preparation of the cell suspension, the HSCs were further isolated using a Nycodenz density gradient. Cell ...
4. Place 20 to 50 μL of the cell suspension (at least enough to cover the well) in each well of the adhesion slides and let cells adhere at room temperature (RT) for 20 min. 5. Continue with the staining procedure. Centrifugation method 1. Assemble the CytoSpin centrifuge's sample ...
Single-cell suspensionSolid tissueTissue digestionDue to its technological advantages, single-cell sequencing has become an increasingly utilized tool to unravel the heterogeneity and complexity of individuals. The preparation of a high-quality single-cell suspension from solid tissues is a critical step ...
Image: A quality single-cell suspension will have high viability, well preserved cell surface antigens, and a low incidence of aggregates or cell debris. Image provided by Agilent Technologies. Jana Soyka, global product manager for sample preparation, cancer research, and MACSmolec...
which needed to be above 85% and single-cell suspensions were adjusted in concentration to 700–1200 cells/μl. After completing the tissue digestion and single-cell suspension preparation, as indicated by the 10×Genomics pre-sequencing needed, we followed by cell sorting, reverse transcription, ...
After performing the second filtration with a cell strainer, the single-cell suspension was stained with fluorescein diacetate (FDA) at 14 mM final concentration for 3 min on ice and the number and activity of cells was calculated with microscope (ZEISS Axioscope A1). The protoplasts with a...
The suspension was then digested utilizing the gentleMACS Octo Dissociator (Miltenyi Biotec, Cat. No. 130-095-937) for 25–30 min using the “37C_h_TDK_3” program for tumor tissue. Dissociated cells were then quenched with ice-cold RPMI 1640 containing 10% fetal bovine serum (FBS; ...
Single-cell suspension preparation Bladder tumors and adjacent normal mucosa were processed immediately after being obtained from bladder cancer patients. Every sample was cut into small pieces (<1 mm in diameter) and then was incubated with 2 ml of trypsin (Gibco, Cat: R001100), 1 ml of ...
In one instance the soleus muscles from two mice (one PdgfraH2BGFP and one PdgfrbGFP) were pooled prior to dissociation for single cell suspension preparation. The tissues were then cut into smaller pieces and incubated in dissociation buffer (Skeletal Muscle Dissociation kit from Miltenyi, ...