Everything counts. But nothing counts like the

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1 Everything counts But nothing counts like the Countess II FL Automated Cell Counter

The new Countess II FL Automated Cell Counter is a benchtop cell assay platform equipped with state-of-the-art optics and image analysis software for rapid assessment of cells in suspension.

2 The new Countess II FL Automated Cell Counter is a benchtop cell assay platform equipped with state-of-the-art optics and image analysis software for rapid assessment of cells in suspension. With three-channel flexibility brightfield and two optional fluorescence channels researchers can count cells, monitor fluorescent protein expression, and measure cell viability. Flexibility choose the fluorescence optical filters for your application Value reusable counting slide reduces consumable costs Precision autofocus designed to improve accuracy and reduce variability Simple easy-to-use touch-screen interface I would describe the new Countess II FL Automated Cell Counter as accurate, consistent, and effortless. Ravi Pathak, PhD Baylor College of Medicine 2 Life Technologies Countess II FL Automated Cell Counter

The advanced counting algorithms of the Countess II FL Automated Cell Counter can identify clear delineations of cell boundaries within clumps of cells, thereby giving precise, accurate cell counts

3 Clumpy cells? Not a problem with the Countess II FL Automated Cell Counter. Whether using automated or manual cell counting methods, obtaining accurate cell counts in samples containing clumpy cells is difficult to achieve. The advanced counting algorithms of the Countess II FL Automated Cell Counter can identify clear delineations of cell boundaries within clumps of cells, thereby giving precise, accurate cell counts even with clumpy samples (Figure 1). Figure 1. Accurate cell counts even with clumpy samples on the Countess II FL Automated Cell Counter. NIH/3T3 cells were mixed 1:1 in suspension with a 0.4% solution of Trypan Blue Stain (Cat. No. T10282), and a 10 μl sample was added to a chamber slide (Cat. No. C10228) 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. Reusable or disposable slides The Countess II FL Automated Cell Counter offers the option of using a reusable slide for counting cells, thereby significantly reducing long-term consumable costs. For those who prefer the convenience of disposable slides, the same slides that are used with the original Countess Automated Cell Counter can also be used with the the Countess II FL Automated Cell Counter (Figure 2). Figure 2. Reusable slide available for the Countess II FL Automated Cell Counter. Precise, accurate counts The Countess II FL Automated Cell Counter contains advanced autofocusing and counting algorithms to quickly and accurately identify and count cells within a population. Upon insertion of the slide, the Countess II FL counter will automatically focus on the cells to avoid user variation associated with manual focusing mechanisms (Figure 3). 3.00E E E E E E+05 Countess II FL Hemocytometer 0.00E+00 NIH/3T3 HeLa HL-60 Figure 3. Precise, accurate cell counts with the Countess II FL Automated Cell Counter. NIH/3T3, HeLa, and HL-60 cells were mixed 1:1 in suspension with a 0.4% solution of Trypan Blue Stain (Cat. No ) and counted using the trypan blue exclusion method. 10 μl were added to a chamber slide (Cat. No. C10228) and inserted into a Countess II FL Automated Cell Counter (blue bar) or hemocytometer and counted manually under a microscope (purple bar). Each bar is an average of 10 counts, with standard deviation as shown. The autofocusing and advanced counting algorithms of the Countess II FL Automated Cell Counter give a more precise, accurate cell count compared to manual counting. Life Technologies Countess II FL Automated Cell Counter 3

Fluorescence counting that fits your experiments The Countess II FL Automated Cell Counter uses the same fluorescent light cubes that power the EVOS fluorescence imaging systems.

With interchangeable light cubes, the Countess II FL Automated Cell Counter was designed to evolve with your changing laboratory needs.

Full spectrum of fluorescence detection With over 15 EVOS light cubes from which to choose, the Countess II FL Automated Cell Counter is not limited to detection of GFP- and RFP-expressing cells

4 Fluorescence counting that fits your experiments 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 was designed to evolve with your changing laboratory needs. As your experiments change, the fluorescence capabilities of the Countess II FL counter can be upgraded with new EVOS light cubes. Full spectrum of fluorescence detection With over 15 EVOS light cubes from which to choose, the Countess II FL Automated Cell Counter is not limited to detection of GFP- and RFP-expressing cells (Table 1). Looking to measure the transfection efficiency of cells transduced with an RFP-expression construct with a blue nuclear counterstain? No problem. Simply add the RFP and DAPI light cubes, and you can effectively count the number of cells expressing the RFP fusion within the total population and accurately determine the transfection efficiency (Figures 4 and 5). Figure 4. Two-color GFP and RFP fluorescent cell counting. 10 µl of HeLa cells transduced with CellLight Histone 2B-RFP (C10595) and stained with NucBlue Live ReadyProbes Reagent were added to a chamber slide (C10228) and inserted into a Countess II FL Automated Cell Counter equipped with DAPI (AMEP4650) and RFP (AMEP4652) light cubes. The segmentation and counting algorithms of the new Countess II FL counter identifies and counts the cells based on their fluorescence properties. In this experiment, 86% of the HeLa cells within the population were transduced with the CellLight Histone 2B-RFP construct. Figure 5. Two-color GFP and DAPI fluorescent cell counting. 10 µl of HeLa cells transduced with CellLight Mitochondria-GFP (C10600) and stained with NucBlue Live ReadyProbes Reagent were added to a chamber slide (C10228) and inserted into a Countess II FL Automated Cell Counter equipped with DAPI (AMEP4650) and GFP (AMEP4651) light cubes. The segmentation and counting algorithms of the new Countess II FL counter identifies and counts 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. 4 Life Technologies Countess II FL Automated Cell Counter

EVOS light cubes for the Countess II FL Automated Cell Counter At the heart of the Countess fluorescence technology lie the proprietary EVOS LED light cubes.

The wide variety of light cubes available provides flexibility for multiplefluorescence research applications, like cell viability analysis (Figure 6). Table 1.

5 EVOS light cubes for the Countess II FL Automated Cell Counter At the heart of the Countess fluorescence technology lie the proprietary EVOS LED light cubes. Each cube contains an LED, illuminating optics, and filters. Light cubes are user interchangeable, auto-configured by the Countess system with plug-and-play capability. The wide variety of light cubes available provides flexibility for multiplefluorescence research applications, like cell viability analysis (Figure 6). Table 1. Common fluorescent light cubes available for the Countess II FL Automated Cell Counter Light cube Excitation (nm) Emission (nm) Common compatible dyes/fluorescent proteins DAPI 357/44 447/60 DAPI, Hoechst, BFP TagBFP 390/18 447/60 TagBFP CFP 445/45 510/42 ECFP, Lucifer Yellow GFP 470/22 510/42 GFP, Alexa Fluor 488, SYBR Green, FITC YFP 500/24 524/27 EYFP, acridine orange (+DNA) RFP 531/40 593/40 Texas Red 585/29 624/40 RFP, Alexa Fluor 546, Alexa Fluor 555, Cy 3, DsRed, Rhodamine Red, dtomato Texas Red, Alexa Fluor 568, Alexa Fluor 594, MitoTracker Red, mcherry Cy 5 628/40 692/40 Cy 5, Alexa Fluor 647, Alexa Fluor 660, DRAQ5 Cy /46 794/16 Cy 5.5 For a complete list of available common and specialty EVOS light cubes, go to lifetechnologies.com/evoslightcubes Figure 6. Cell viability. Heat-treated HL-60 cells were combined with non-treated cells at approximately a 15:85 ratio. To assess cell viability of the population, cells were stained with a combination of NucBlue Live and NucGreen Dead 488 stains. 10μL of cells were added to a chamber slide and inserted into a Countess II FL Automated Cell Counter equipped with DAPI (AMEP4650) and GFP (AMEP4651) light cubes. As expected, 14% of the HL-60 cell population were dead as indicated by positive NucGreen Dead 488 (green) staining. Life Technologies Countess II FL Automated Cell Counter 5

6 Table 2. Cell lines validated on the Countess II FL Automated Cell Counter Cell type Animal Organ A431 Human Skin Adipocytes Human Adipose-derived stem cells Aortic smooth muscle Human Smooth muscle CHO-M1WT2 Chinese hamster Ovary CHSE Chinook salmon Embryo COLO-205 Human Colon COS-7 African monkey Kidney HEK-293 Human Kidney HeLa Human Cervix HepG2 Human Liver HL-60 Human Blood J774A.1 Mouse Blood Jurkat Human Blood K562 Human Bone marrow MCF7 Human Breast MRC-5 Human Lung NIH/3T3 Mouse Embryo PC-12 Rat Adrenal gland Pulmonary artery endothelial cells Human Blood vessel Pulmonary artery smooth muscle Human Smooth muscle SF-21 Insect Ovary U266 Human Blood U-2 OS Human Bone Umbilical vein endothelial cells Human Blood vessel 6 Life Technologies Countess II FL Automated Cell Counter

7 Table 3. Countess II FL Automated Cell Counter technical specifications Technical specifications Optics Physical characteristics Processing time: Sample concentration range: Particle/cell diameter range: Required sample volume: Firmware: USB drive: Optics: Camera: Instrument type: Instrument dimensions: Weight: Operating power: Frequency: Electrical input: Installation site: Operating temperature: Operating humidity: <10 seconds to cells/ml μm 10 μl Countess Automated Cell Counting Platform Software 3 channels (brightfield and 2 slots for EVOS LED light cubes) 5 megapixels, 2.5x optical magnification Benchtop cell counter and suspension cell based assay platform 9" (W) 5½" (D) 9" (H) mm (W) X mm (D) x mm (H) 8 lb VAC, 0.58 A max 50/60 Hz 12 VDC, 2 A Indoor use only, Class A environments (i.e., nonresidential or light industrial); Pollution degree C <80% (noncondensing) Material: Glass Reusable analysis slide Disposable analysis slide Dimensions: Chamber volume: Material: Dimensions: Chamber volume: Slide: 32 mm (W) 76 mm (D) 3.9 mm (H) Coverslip: 20 mm (W) x 26 mm (L) x 0.5 mm (H) 10 μl Poly(methyl methacrylate) (PMMA) Slide: 25 mm (W) 75 mm (D) 1.7 mm (H) 10 μl Life Technologies Countess II FL Automated Cell Counter 7

8 Need a simple, brightfield cell counter? Check out the Countess II Automated Cell Counter. Feature Countess II Countess II FL Autofocus Touch-screen Brightfield counting Trypan blue counting Disposable slide Reusable slide Fluorescence compatible/upgradable Uses EVOS light cubes Find out more at lifetechnologies.com/countessii Ordering information Product Quantity Cat. No. Countess II FL Automated Cell Counter 1 AMQAF1000 Countess II Automated Cell Counter 1 AMQAX1000 Countess Cell Counting Chamber Slides 50 C10228 Countess II FL Reusable Slide 1 A25750 Countess II FL Reusable Slide Holder 1 AMEP4746 EVOS Light Cube, DAPI 1 AMEP4650 EVOS Light Cube, TagBFP 1 AMEP4668 EVOS Light Cube, YFP 1 AMEP4654 EVOS Light Cube, GFP 1 AMEP4651 EVOS Light Cube, RFP 1 AMEP4652 EVOS Light Cube, Texas Red 1 AMEP4655 EVOS Light Cube, Cy 5 1 AMEP4656 EVOS Light Cube, Cy AMEP4673 Find out more at lifetechnologies.com/countessii For Research Use Only. Not for use in diagnostic procedures Thermo Fisher Scientific Inc. All rights reserved. All trademarks are the property of Thermo Fisher Scientific and its subsidiaries unless otherwise specified. Cy is a registered trademark of GE Healthcare. DRAQ5 is a registered trademark of Biostatus Limited. CO

Everything counts. But nothing counts like Countess II automated cell counters. Fast Accurate Affordable

Everything counts. But nothing counts like Countess II automated cell counters. Fast Accurate Affordable Everything counts But nothing counts like Countess II automated cell counters Fast Accurate Affordable Countess II automated cell counters Advanced technology at an affordable price Precise, accurate counts