Introducing a Three-Channel Cell Counter for Your Benchtop

The new and improved Countess® II FL Automated Cell Counter

(See a list of the products featured in this article.)

Since its introduction in 2008, the Countess® Automated Cell Counter has revolutionized the way that researchers determine cell number by removing the tedium of manually counting cells. The original Countess® cell counter uses a sophisticated image analysis algorithm that identifies objects, classifies cells by roundness and size, and then distinguishes live cells from dead cells by their staining patterns. The new Countess® II FL Automated Cell Counter (Figure 1) has been reworked from the inside out, with faster cell analysis, more detection options, and better user controls, all to provide researchers with the ability to configure the instrument to fit their specific cell counting needs.

The Countess® II FL Automated Cell Counter is a three-channel (brightfield and two optional fluorescence channels) benchtop assay platform with state-of-the-art optics and image analysis software to automate cell counting. This instrument allows researchers to analyze cells in suspension for a range of characteristics, including cell viability (live, dead, and total cell counts), cell cycle phase, apoptotic stage, and fluorescent protein expression. For a typical cell count, the Countess® II FL Automated Cell Counter takes less than 10 seconds per sample, using autofocusing to enable fast, accurate counts of cells in a population. The cell counting assay is compatible with a wide variety of eukaryotic cells—including clumpy samples (Figure 2)—using the standard trypan blue exclusion method to determine cell viability in the brightfield channel.

In addition to the brightfield channel, the Countess® II FL Automated Cell Counter features two user-defined fluorescence channels. Along with these enhanced features, the Countess® II FL Automated Cell Counter offers an intuitive user interface as well as the option to save cell count data and generate a report that can then be transferred to the USB drive supplied with the instrument.

Figure 1. The Countess® II FL Automated Cell Counter and light cubes.

Figure 2. Accurate cell counts—even with clumpy samples—on the Countess® II FL Automated Cell Counter. NIH/3T3 cells in suspension were mixed 1:1 with a 0.4% solution of Trypan Blue Stain, and a 10 μL sample was added to a chamber slide and inserted into a Countess® II FL Automated Cell Counter. The segmentation and counting algorithms of the Countess® II FL counter are able to resolve clumps of cells and score them as live or dead using the trypan blue exclusion method. In this sample, 3.09 x 106 live cells (which excluded trypan blue stain) and 5.86 x 103 dead cells (which accumulated trypan blue stain) were counted.

Full spectrum of fluorescence detection: Now and in the future

With more than 15 EVOS® light cubes to choose from, the Countess® II FL Automated Cell Counter is not limited to detection of GFP- and RFP-expressing cells. Looking to measure the transfection efficiency of an mCherry fluorescent protein expression vector introduced into cells? No problem. Simply insert the Texas Red® light cube, and you can effectively count the number of cells expressing the mCherry fusion protein and accurately determine the transfection efficiency. Figure 3 shows two-color fluorescence counting of HeLa cells transduced with CellLight® Mitochondria-GFP, stained with NucBlue® Live ReadyProbes® Reagent, and analyzed on a Countess® II FL Automated Cell Counter equipped with GFP and DAPI light cubes.

The Countess® II FL Automated Cell Counter uses the same fluorescent light cubes that power the EVOS® fluorescence imaging systems. EVOS® light cubes integrate LEDs and high-performance filters in a novel illumination system that delivers precise control, minimal maintenance, and exceptional reliability. With interchangeable light cubes, the Countess® II FL Automated Cell Counter is designed to evolve with your changing laboratory needs. As your experiments demand, the fluorescence capabilities of the Countess® II FL counter can be upgraded with new EVOS® light cubes. The full line of EVOS® light cubes can be found at lifetechnologies.com/evoslightcubes. Or hang the EVOS® Light Cube poster in a convenient spot in your lab so you can see your options at a glance.

Figure 3. Two-color fluorescent cell counting on the Countess® II FL Automated Cell Counter. A 10 μL sample of HeLa cells transduced with CellLight® Mitochondria-GFP and stained with NucBlue® Live ReadyProbes® Reagent was added to a chamber slide and inserted into a Countess® II FL Automated Cell Counter equipped with a DAPI EVOS® Light Cube and a GFP EVOS® Light Cube. The segmentation and counting algorithms of the Countess® II FL counter identify and count the cells based on their fluorescence properties. In this experiment, 72% of the HeLa cells within the population were transduced with the CellLight® Mitochondria-GFP construct.

Reusable glass slide saves you money

Mounting costs of disposable slides and tips have deterred many labs from finding alternatives to the daily chore of manually counting cells. The Countess® II FL Automated Cell Counter is designed to work with a reusable glass slide (Figure 4) that significantly reduces the long-term costs of consumables used for automated counting. For researchers who want the convenience of disposable slides, the Countess® II FL Automated Cell Counter can also use the same slides developed for the original Countess® cell counter. The Countess® cell counting slides contain two enclosed sample chambers, allowing you to measure two different samples or perform replicates of the same sample. Cell counting occurs in the central location of each counting chamber, and the volume counted is 0.4 μL, which is equivalent to counting four 1 mm x 1 mm squares in a standard hemocytometer.

Figure 4. Precise cell counts with the Countess® II FL Automated Cell Counter. NIH/3T3, HeLa, and HL-60 cells in suspension were mixed 1:1 with a 0.4% solution of Trypan Blue Stain and counted using the trypan blue exclusion method. A 10 μL sample was added to a chamber slide and inserted into a Countess® II FL Automated Cell Counter (blue bars) or counted manually under a microscope using a hemocytometer (purple bars). Each bar is an average of 10 counts, with standard deviation as shown. The autofocusing and advanced counting algorithms of the Countess® II FL counter give more precise cell counts than those derived from manual counting.

Bring automated cell counting to your benchtop

The Countess® II FL Automated Cell Counter is designed to adapt to your ever-changing cell counting needs. With two user-defined fluorescence channels and the option for a reusable slide, nothing counts like the Countess® II FL Automated Cell Counter.