Background

Isolation and cryopreservation of peripheral blood mononuclear cells (PBMC) is common practice in clinical studies that employ cellular immune assays. Cryopreservation allows for batching of samples, which is convenient and improves the comparability of data. Cryopreservation also allows cells to be stored for unknown future purposes. Because erythrocytes and granulocytes are much more fragile to freezing and thawing, PBMC isolation is a common prerequisite to cryopreservation of blood cells; and the most common method for PBMC isolation is density gradient centrifugation using Ficoll-Hypaque, a high molecular weight carbohydrate solution.

Cellular Preparation Tubes (CPTs), which contain Ficoll-Hypaque, simplify the standard procedure for density gradient centrifugation in two ways: (1) blood is collected into the same tube that is then used to isolate the PBMC; and (2) the tube is pre-loaded with Ficoll-Hypaque, which is separated by a gel plug so that it is not disturbed by the entry of blood into the tube. After blood draw, the tubes are centrifuged, and the PBMCs and plasma become separated from the erythrocytes and granulocytes by the gel plug (see Figure 1). This allows the spun tubes to be shipped, maintaining the PBMC in an isolated environment from the erythrocytes and granulocytes, which may improve their viability and function.

CPTs are ideal for use in studies which collect whole blood across multiple sites and ship to a central processing laboratory (Ruitenberg et al., 2006); this reduces variability in PBMC isolation between technicians. Studies have found no significant difference in PBMCs isolated using the CPT system or by traditional layover methods via density gradient separation (Corkum et al., 2015; Ruitenberg et al., 2006). While material cost can be high, CPTs reduce the length of processing time in addition to decreasing inconsistency between operators; both of these are key to reducing cost and increasing sample quality and consistency. Furthermore, when used with studies or sites which ship blood samples overnight, PBMCs from CPTs have a higher purity and less infiltrate from other (contaminating) cell types, such as red blood cells, in contrast to samples shipped in standard blood collection tubes over a 24-48 h period (Schlenke et al., 1998).


Figure 1. Empty CPT (left), after blood draw (middle), and after centrifugation (right). Location of gel plug and sample layers after centrifugation are shown.