Experimental Insights! Pre-treatment Tips to Prevent Column Clogged.

The preparation of high-quality single-cell suspension is the key to carry out downstream separation experiments. High viability, excellent dispersion, no debris, and other factors are the criteria used to evaluate the quality of a single-cell suspension.

Enzymolysis, as one of the common methods for tissue dissociation, involves the digestion of tissue into individual cells through corresponding chemical reactions facilitated by enzymes. Collagenase, papain, and various other enzymes are commonly employed for this purpose, and tissue digestion can be accomplished using either a single enzyme or a combination of enzymes.

The sample can be filtered using a cell strainer to further remove debris, erythrocytes, etc.

The cell suspension should be counted without exceeding the maximum recommended capacity of the cell separation columns.

Prior to adding the cell suspension, carefully inspect the liquid for the presence of any bubbles. If bubbles are detected, they can be eliminated by gently agitating the solution. It is crucial to add cell suspension slowly and cautiously to prevent the formation of bubbles.

Tips to improve purity and viability of cell separation experiments.

Both the antibodies and microbeads solutions are homogeneous colloids. When pipetting, avoid shaking to prevent the formation of bubbles, which can cause errors in volume measurement.

To ensure optimal binding efficiency, the cell suspension should be gently mixed with microbeads or antibodies using a pipette, taking care to avoid the formation of bubbles. When dealing with a higher cell volume, it is advisable to gently shake the sample during the incubation process to prevent cellular sedimentation, which may otherwise compromise binding efficiency.

Optimal incubation should be conducted in a refrigerator set between 2 and 8°C. Failure to incubate at a lower temperature may lead to decreased cell activity.

Place the sample solution at room temperature before passing through the column. Failure to follow this step may lead to excessive bubbles in the cell separation columns and compromise the desired purity levels.