Scepter™ Automated Cell Counter from Millipore

Much of our research of late has involved cell-based assays such as ADCC (antibody-dependent, cell-mediated cytotoxicity) and migration. Since setting up and evaluating these assays involves precise determinations of cell number, we are always on the lookout for automated cell counters that eliminate the need for tedious and inaccurate manual counting methods. Over the past few years Bio-Rad, Invitrogen, Nexcelom, and Millipore have introduced the TC10™, Countess®, Cellometer®, and Scepter™ automated cell counters, respectively. While the Bio-Rad, Invitrogen, and Nexcelom instruments rely on the identification and counting of cells by scanning, the Scepter uses the same technology as that used by Coulter Counters. The Coulter technology is based on the fact that the electrical resistance between two electrolyte-filled compartments changes when a particle, such as a cell, traverses a pore connecting the compartments. This change in resistance can be detected and converted into cell counts. Using microfluidics, Millipore has managed to miniaturize this technology into a hand-held device the size of a pipettor.

Over a period of a few weeks, our very accommodating Millipore sales rep brought a Scepter to our lab for evaluation at least three different times. We tested it with cultured lung cancer cells, freshly isolated PBMCs, and CD34+ hematopoietic progenitor cells isolated from umbilical cord blood. Using the Scepter is very easy: You first attach a disposable microfluidic tip to the Scepter, depress the plunger, submerge the tip in a small volume of diluted cells, release the plunger to allow the cell suspension to be drawn into the tip, and wait a few seconds for the count histogram to appear on the display screen. The histogram displays cell size, in pL or diameter, and total cell count. The Scepter will store up to 70 histograms, which can be downloaded to a PC at any time and analyzed further using the included software. Cells diluted to approximately 50,000 to 500,000 cells/ml can be counted in the Scepter. In practice, we have observed accurate cell counts down to about 10,000 cells/ml.

One advantage of being able to see the range of particle sizes with the histogram is that very small cellular debris as well as enlarged, and probably dead, cells can be identified and eliminated from the count. Since both lower and upper size limits can be set by the user after the cell count, counts with or without aberrantly sized particles can be determined easily. Once the limits are set, the user can choose to apply these limits to subsequent counts. One downside, however, is that dead cells are only inferred from cell diameter. If you need an accurate estimate of live/dead cells, you will need to check them visually with trypan blue.

The recommended cell size range for counting in the Scepter is 8 – 25 um diameter. Since some PBMCs and progenitor cells are near the lower limit of this range, the histogram abuts the y-axis, suggesting that some cells might be missed. A clever solution suggested by the Millipore rep is to dilute the cells into 60-65% PBS prior to counting. This causes sufficient osmotic swelling to shift the average cell diameter to above the 8 um lower limit such that all cells get counted. While we have verified that this method works very well, Millipore will soon offer a tip with a smaller lower limit of detection. This should allow counting of all cell types in the Scepter without osmotic manipulation.