Coulter particle counter

Manufactured by Beckman Coulter

Sourced in United States

The Coulter Particle Counter is a laboratory instrument used for counting and sizing particles suspended in a fluid. It operates on the principle of electrical sensing zone technique, which measures changes in electrical resistance as particles pass through a small aperture. The device provides accurate and reliable data on the number and size distribution of particles within a sample.

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15 protocols using coulter particle counter

Evaluating Fibroblast Proliferation Induced by NE

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Human dermal fibroblasts (5 × 10⁵ cells per well) were plated in a 6 cm² dish and starved in serum free medium overnight. Cells were treated with or without NE (10 μM). After 48 hours incubation at 37 °C, cells were counted by Coulter particle Counter (Beckman coulter, Brea, CA).

Uehara A., Motegi S.I., Yamada K., Uchiyama A., Perera B., Toki S., Ogino S., Yokoyama Y., Takeuchi Y, & Ishikawa O. (2016). Mechanistic insight into the norepinephrine-induced fibrosis in systemic sclerosis. Scientific Reports, 6, 34012.

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LOX Expression and Cell Growth

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To study the effect of LOX re-expression on cell growth, we used Odc cells carrying the tetracycline-inducible pLRT-LOX or empty vector. The cells were plated in triplicates in the absence or presence of doxycycline (1 μg/ml) and the total numbers of cells were counted with a Coulter particle counter (Beckman Coulter, Fullerton, CA, USA) after 3 and 5 days (medium changed after 3 days). To study the effect of ΒAPN on cell growth of Odc and WM793 cells, the cells were cultured in triplicates with 0, 250, or 500 μM BAPN and counted after 3 and 5 days. The medium and ΒAPN were changed after 3 days. In addition, the cell numbers of WM793 cells transduced with control shRNA, LOX shRNA, or two different LOXL2 shRNAs were counted 10 days after transduction with the shRNA lentiviral transduction particles.

Kielosto M., Eriksson J., Nummela P., Yin M, & Hölttä E. (2018). Divergent roles of lysyl oxidase family members in ornithine decarboxylase- and RAS-transformed mouse fibroblasts and human melanoma cells. Oncotarget, 9(102), 37733-37752.

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Lentiviral Transduction and Cell Growth

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Cells were infected with either NT control or target lentivirus as previously described. Cells were plated at 2x10⁴ cells/well in 12-well plates (Midwest Scientific) in triplicate. After 5 days, cell number was established using a Coulter Particle Counter (Beckman). Puromycin selection was maintained throughout the course of the assay.

von Roemeling C.A., Marlow L.A., Radisky D.C., Rohl A., Larsen H.E., Wei J., Sasinowska H., Zhu H., Drake R., Sasinowski M., Tun H.W, & Copland J.A. (2014). Functional genomics identifies novel genes essential for clear cell renal cell carcinoma tumor cell proliferation and migration. Oncotarget, 5(14), 5320-5334.

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Evaluating Synergistic Dose Combinations

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For dose out curves and lentiviral infected clones, cells were treated with belinostat, vorinostat, romidepsin or paclitaxel at the indicated doses and were counted on a Coulter Particle Counter (Beckman, Brea, CA). When applicable, IC₅₀ was determined via extrapolation of 50% growth on log scale to the corresponding drug concentration. Using the ratio of the IC₅₀, the proportion of each compound needed in a combination dose was calculated. Experiments were then carried out using HDACi, paclitaxel and a fixed ratio combination of both at the indicated doses in clear-bottom black plates (Costar, Corning, NY) and analyzed using the CyQUANT proliferation assay kit (Invitrogen) as per manufacturer’s protocol for relative fluorescence units. Drug interactions were analyzed using CalcuSyn® (Biosoft, Cambridge, UK). Determination of synergy, additivity or antagonism was based on the multiple drug effect equation of Chou and Talalay and was quantified by the combination index (CI). CI = 1 indicates an additive effect, < 1 is synergy and > 1 is antagonism (Chou and Talalay 1984 (link)).

Marlow L.A., Bok I., Smallridge R.C, & Copland J.A. (2015). RhoB upregulation leads to either apoptosis or cytostasis through differential target selection. Endocrine-related cancer, 22(5), 777-792.

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Platelet Aggregation Assay Protocol

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Washed platelets were used to conduct light transmission aggregometry. Platelet concentrations were assessed using a Coulter Particle Counter (Beckman Coulter, CA), and samples adjusted to 2.5 × 10⁸ platelets/ml using PBS^−/− prior to platelet aggregometry. Studies were conducted on 225 μl of washed platelets. Immediately prior to aggregation, 4 μl of 100 mM CaCl₂ and 1 μl of 100 mM MgCl₂ were added to each sample. Platelet aggregation in response to calf thymus histone (CTH, 0.75 mg/ml), collagen (2 μg/ml), or saline control was measured in the washed platelets at 37°C and at 1200 rpm in a Bio/Data PAP-8 aggregometer (Bio/Data Corporation, PA).

Courson J.A., Lam F.W., Langlois K.W, & Rumbaut R.E. (2022). Histone-stimulated platelet adhesion to mouse cremaster venules in vivo is dependent on von Willebrand factor. Microcirculation (New York, N.Y. : 1994), 29(8), e12782.

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Cell Culture Conditions for RUNX1 Models

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HeLa cells were cultured in DMEM medium (Thermo Fisher, Waltham, MA) supplemented with 10% FBS (Thermo Fisher). 32D cells expressing wild type RUNX1 and GCSF receptor [17 (link)] and 32D cells carrying the RUNX1-MTG8 fusion protein (32D-RM8) [13 (link)], and derived cell lines were cultured in RPMI 1640 (Thermo Fisher) medium supplemented with 10% heat-inactivated (HI) fetal bovine serum (FBS) (Thermo Fisher) and 1 ng/ml murine IL-3 (Peprotech, Inc., Rocky Hill, NJ). Cells were counted daily with a Coulter Particle Counter (Beckman Coulter, Brea, CA) and diluted to a density of 2 × 10⁵ cells/ml daily.

Rossetti S., Anauo M.J, & Sacchi N. (2017). MiR-221-regulated KIT level by wild type or leukemia mutant RUNX1: a determinant of single myeloblast fate decisions that – collectively – drives or hinders granulopoiesis. Oncotarget, 8(49), 85783-85793.

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Cell Proliferation on ECM Coatings

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Ten thousand (786-O and Caki-2) or 20,000 (Renca) cells in 500 μl of complete medium were cultured for 72 h in a 24 well plate coated or not with the different ECMs. Cells were trypsinized and counted using a Coulter Particle Counter (Beckman Coulter France).

Majo S., Courtois S., Souleyreau W., Bikfalvi A, & Auguste P. (2020). Impact of Extracellular Matrix Components to Renal Cell Carcinoma Behavior. Frontiers in Oncology, 10, 625.

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Isolation of Peripheral Blood Mononuclear Cells

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Peripheral blood mononuclear cells (PBMCs) were isolated from blood of healthy volunteers by density gradient centrifugation using Ficoll-plaque (GE Healthcare, Diegem, Belgium). PBMCs were washed in cold phosphate buffered saline (PBS, Braun Melsungen, Germany) and resuspended in Roswell Park Memorial Institute (RPMI) 1640 Dutch modification culture medium (Life Technologies, Carlsbad, California, USA) supplemented with gentamycin 50 µg/mL, pyruvate 1 mM, glutamax 2 mM and counted using a Coulter particle counter (Beckman Coulter, Fullerton, California, USA). Monocytes were isolated within the PBMC fraction using Percoll as described previously.21 (link)

Rabold K., Aschenbrenner A., Thiele C., Boahen C.K., Schiltmans A., Smit J.W., Schultze J.L., Netea M.G., Adema G.J, & Netea-Maier R.T. (2020). Enhanced lipid biosynthesis in human tumor-induced macrophages contributes to their protumoral characteristics. Journal for Immunotherapy of Cancer, 8(2), e000638.

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PBMC Isolation and Cryopreservation

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PBMCs were obtained following gradient centrifugal separation of peripheral blood using Cell Preparation Tubes (CPT; BD Biosciences, Franklin Lakes, NJ, USA) for 30 minutes at 1700 x g. Plasma was separated, aliquoted, and stored at -20°C for antibody assays. The mononuclear layer was washed in PBS prior to counting on a Coulter particle counter (Beckman Coulter, Brea, CA, USA). PBMCs were either directly stained for flow cytometry (initial visit samples), or frozen slowly to -80°C in FBS and DMSO for short-term storage (3-month visit samples). Flow cytometry panel was validated using a sample of fresh vs. frozen PBMCs to ensure comparability in target detection.

Newell K.L., Clemmer D.C., Cox J.B., Kayode Y.I., Zoccoli-Rodriguez V., Taylor H.E., Endy T.P., Wilmore J.R, & Winslow G.M. (2021). Switched and unswitched memory B cells detected during SARS-CoV-2 convalescence correlate with limited symptom duration. PLoS ONE, 16(1), e0244855.

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Cell Viability Assessment Protocols

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Cell counting was performed using a Coulter Particle Counter (Beckman-Coulter). For ATP bioluminescence experiments, cells were plated at a density of 1,500 (HARA, HCC1438, NCI-H2052, PATU-8902) or 3,000 (A375, A549, KYSE-70, MDA-MB-157, NCI-H460, NCI-H1299, NCI-H1437, HCC366, NCI-H1650) cells per well in 96 well assay plates (Corning). ATP bioluminescence was assessed at 12 days after gene KO with the CellTiter-Glo Luminescent Cell Viability Assay (Promega). For crystal violet staining assays, cells were plated at a density of 10,000 or 20,000 cells per well in 12 well tissue culture plates. Once the control cells grew to near confluency, each well was washed twice with ice-cold PBS, fixed on ice with ice-cold methanol for 10 min, stained with 0.5% crystal violet solution (made in 25% methanol) for 10 min at room temperature, and washed at least four times with water. All cell viability assays were performed in at least technical and biological triplicates.

Zou T., Zhou M., Gupta A., Zhuang P., Fishbein A.R., Wei H.Y., Capcha-Rodriguez D., Zhang Z., Cherniack A.D, & Meyerson M. (2024). XRN1 deletion induces PKR-dependent cell lethality in interferon-activated cancer cells. Cell reports, 43(2), 113600.

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