Flow Cytometry

Hello Flow-ers, Flowbos, Cytometrians, or just fellow flow cytometry lovers. Welcome to our Flow Cytometry division! We’ve been in beads a long time, and are proud to continue the tradition of standardization in flow cytometry that our predecessor, Flow Cytometry Standards Corporation began. We offer a broad range of products that support instrument validation and QC, and assay and multi-site standardization.

Flow Cytometry Products

Instrument Set-Up

In flow cytometry, consistent instrument set-up and standardization are essential for achieving reliable results and generating meaningful data. Our microsphere-based tools for set-up and standardization can serve a significant role in supporting flow cytometry programs, and establishing a framework for the comparison of data from different instruments, laboratories, and over time.

QUANTUM™ QC, 8 PEAK BEADS

Quantum™ QC is an 8 peak multi-intensity, multi-fluorescent standard that may be used to set up the window of analysis for all detectors by positioning a specific peak at a target channel value. See PDS 725 & 725A for histograms for all detectors.

Catalog Number	Description
725	Quantum™ QC, 8 Peak Beads

REFERENCES

Purvis N, Stelzer G. (1998) Multi-platform, multi-site instrumentation and reagent standardization. Cytometry; 33(2):156-65.

FULL SPECTRUM™

Full Spectrum™ Multi-Color Fluorescence Reference Standard is a highly uniform microbead product that fluoresces over the visible spectrum to provide reference positions for initial daily QC or instrument set-up.

As part of a QC program, fluorophore-labeled microspheres offer a convenient means to check general instrument status and monitor stability over time. When beads are run, the channel values for pertinent fluorescence detectors are recorded, and data are monitored to identify outliers and trends. This type of check can alert the user to problems with the optical and fluidic systems (e.g. diminishing laser power or obstruction/leakage) and the effect of environmental factors, such as temperature, humidity, and vibration on instrument performance.

As a tool for instrument set-up, a reference peak may be positioned for each detector to achieve initial instrument settings.

Catalog Number	Description
885	Full Spectrum™

ULTRA RAINBOW FLUORESCENT PARTICLES

Ultra Rainbow Fluorescent Particles (by Spherotech™) are internally labeled with multiple fluorophores, enabling excitation at any wavelength from 365-650nm. Ultra Rainbow Fluorescent Particles may be used for general QC purposes for 3-9 color instruments. Ultra Rainbow Fluorescent Particles are available in three amounts: A (20 tests), B (100 tests), and C (280 tests).

Catalog Number	Description
610	Ultra Rainbow Fluorescent Particles, ~3.8µm (3.61-3.99)
611	Ultra Rainbow Fluorescent Particles ~10.2µm (8.1-12.0)

SMALL BEAD CALIBRATION KITS

Current applications in flow cytometry extend far beyond traditional lymphocyte immunophenotyping, with some involving the analysis of very small particles such as platelet- and endothelial-derived microparticles or microbial species. Our fluorescent Small Bead Calibration Kits allow operators to verify the resolution capabilities of the flow cytometer, and to establish appropriate instrument settings for the analyses.

Catalog Number	Description
834	Nanobead Calibration Kit – 50nm, 100nm
832	Submicron Bead Calibration Kit – 0.2µm, 0.5µm, 0.8µm
833	Micron Bead Calibration Kit – 1.0µm, 3.0µm, 6.0µm

Instrument QC

Fluorophore-labeled microspheres offer a convenient means to check general instrument status and monitor stability over time. When beads are run, the median channel values for pertinent fluorescence detectors are recorded, and data are monitored to identify outliers and trends. This type of check can alert the user to problems with the optical and fluidic systems (e.g. diminishing laser power or obstruction / leakage) and the effect of environmental factors such as temperature, humidity, and vibration on instrument performance.

QUANTUM™ QC, 8 PEAK BEADS

Quantum™ QC is an 8 peak multi-intensity, multi-fluorescent standard that is intended for use as an in-depth tool for daily cytometer QC, and is appropriate for use with all lasers and detectors. It may be used to determine detection thresholds, understand resolution, and assess and track linearity of detectors. It can aid in providing confidence that the system is suitable for use, or alert operators to potential problems before samples are run. Quantum™ QC may also be used for instrument set-up to achieve standardized PMT settings and define the window of analysis for relevant detectors. Quantum™ QC is comprised of a blank and 7 (mixed) intensity populations. Quantum QC is used with a QuickCal-Linearity template that is downloaded from our website with an Access Code provided with the product at the time of shipping.

Catalog Number	Description
725	Quantum™ QC, 8 Peak Beads

FULL SPECTRUM™

Full Spectrum™ microspheres provide a convenient means for performing the initial daily QC check on 3+ color instruments. These ~7-9µm microspheres are internally labeled with multiple fluorophores for use with common excitation and detection wavelengths within the visible spectrum. Full Spectrum™ are available in three amounts: A (20 tests), B (100 tests), and C (280 tests). See PDS 885 for representative histograms.

Catalog Number	Description
885	Full Spectrum™

RIGHT REFERENCE STANDARD™

Each standard consists of a population of microspheres surface labeled with a single fluorochrome at a high intensity level. The Right Reference Standard™ microspheres approximate the size of human lymphocytes (~7-9µm). With their narrow fluorescence CVs, Right Reference Standard™ products are suitable for alignment purposes. These standards may also be used to QC a specific path of the optical system (laser/filter/PMT), and to establish a unified fluorescence range for a particular detector.

Catalog Number	Description
512	Right Reference Standard™ Fluorescein High Level
515	Right Reference Standard™ Phycoerthrin High Level
518	Right Reference Standard™ PE-Cy™5 High Level
521	Right Reference Standard™ APC High Level

TIME DELAY CALIBRATION STANDARD

Bangs’ Time Delay Calibration Standard is intended for use in assessing the delay between blue and red lasers. It features ~6µm microspheres dyed with a fluorophore that is excited with 488nm or 635nm excitation, and exhibits red/far-red emission.

Catalog Number	Description
830	Time Delay Calibration Standard

FLUORESCENCE REFERENCE STANDARDS

Single-color Fluorescence Reference Standards are labeled with specific fluorochromes to exhibit the same spectral characteristics as labeled cells. They may be used to QC a specific path of the optical system, to optimize filter sets for fluorophores and to establish a test-specific Target Channel Value for instrument set-up. See flow cytometry fluorophore reference chart here.

Catalog Number	Description
890	Certified Blank™
897	Acridine Orange
886	Alexa Fluor® 488
887	Alexa Fluor® 647
901	Allophycocyanine (APC)
898	Chlorophyll
895	Cy™5
906	DAPI
913	Far-Out Red
891	Fluorescein (FITC),
894	Hoechst 33342
916	Pacific Blue™
908	PE-Cy™5
889	PE-Cy™7
909	PE-TR
892	Propidium Iodide
899	R-Phycoerythrin (PE)
905	T.M. Rhodamine
893	Texas Red® (TR)
915	Violet Laser

Compensation Controls

Bangs offers a broad portfolio of standards for color compensation. Our offerings include beads that may be labeled with the antibody conjugate or amine-reactive dye of choice, as well as reference standards pre-labeled with common flurophores.

FLOW CYTOMETRY ANTIBODY BINDING BEADS

Single population Protein A or Protein G microspheres are suitable for labeling with conjugated antibodies from a range of hosts. Labeled microspheres may be used as single-population reference standards or in conjunction with an unlabeled population for compensation purposes.

Catalog Number	Description
553	Flow Cytometry Protein A Antibody Binding Beads

REFERENCES

Musil, J, Kutinova, L, Zurkova, K, Hainz, P, Babiarova, K, Krystofova, J, & Nemeckova, S. (2014) Antitumor activity and immunogenicity of recombinant vaccinia virus expressing HPV 16 E7 protein SigE7LAMP is enhanced by high-level coexpression of IGFBP-3. Cancer gene therapy, 21(3), 115.

SIMPLY CELLULAR^® COMPENSATION STANDARD

The Simply Cellular^® Compensation Standard includes a mixed population of low- and high-binding antibody-capture beads that are ~7-9µm in diameter. Users label aliquots with fluorescent antibody conjugates of interest to establish suitable compensation settings for a specific analysis.

Catalog Number	Description
550	Simply Cellular® Compensation Standard (anti-Mouse IgG)
556	Simply Cellular® Compensation Standard – High (anti-Mouse IgG)
551	Simply Cellular® Compensation Standard (anti-Rat IgG)
552	Simply Cellular® Compensation Standard (anti-Human IgG)

REFERENCES

Wang J-C E, Kobie JJ, Zhang L, Cochran M, Mosmann TR, Ritchlin CT, Quataert SA. (2009) An 11-color flow cytometric assay for identifying, phenotyping, and assessing endocytic ability of peripheral blood dendritic cell subsets in a single platform. J Immunol Methods; 341:106-116.
Scheible K, Secor-Socha S, Wightman T, Wang H, Mariani T.J., Topham DJ,…Quataert S (2012) Stability of T Cell Phenotype and Functional Assays Following Heparinized Umbilical Cord Blood Collection. Cytometry A 81(11):937–949

SIMPLY CELLULAR^® ANTI-MOUSE FOR VIOLET LASER

The Simply Cellular^® anti-Mouse for Violet Laser standard features microspheres comprised of a proprietary matrix that exhibits low autofluorescence with violet excitation. Beads are suitable for labeling with mouse mAbs conjugated with violet fluorochromes, and for use as a compensation or general reference standard for detectors off of the violet laser. Beads are also suitable for use with other fluorochromes / detectors, e.g. 488nm, 633nm. The Simply Cellular^® anti-Mouse for Violet Laser standard is supplied as 2 populations: 1 blank and 1 high-binding anti-Mouse IgG (Fc-specific) population, each are supplied in aqueous suspension containing ProClin^®.

Catalog Number	Description
835	Simply Cellular® anti-Mouse for Violet Laser

VIABILITY DYE COMPENSATION STANDARDS

Viability Dye Compensation Standards are suitable for labeling with LIVE/DEAD^® stains or other amine-reactive dyes to generate compensation standards for flow cytometric analyses. Beads are not suitable for labeling with DNA stains such as propidium iodide, DAPI, or SYTOX^®, and users should contact us for discussion if uncertain as to the compatibility of a specific dye or stain.

Catalog Number	Description
450	Viability Dye Compensation Standard, 4µm
451	Viability Dye Compensation Standard, 8µm

REFERENCES

Perfetto SP, Chattopadhyay PK, Lamoreaux L, Nguyen R, Ambrozak D, Koup RA, Roederer M. (2006). Amine reactive dyes: an effective tool to discriminate live and dead cells in polychromatic flow cytometry. J Immunol Methods; 313(1-2):199-208.
Perfetto SP, Chattopadhyay PK, Lamoreaux L, Nguyen R, Amrozak D, Koup RA, Roederer M. (2010) Amine-reactive dyes for dead cell discrimination in fixed samples. Curr Protoco Cytom; Jul; CHAPTER: Unit-9.34. (SIMPLX Amine active beads [Cat. 450, 451]).,
Lamoreaux, L, Roederer, M, Koup, R (2006) Intracellular cytokine optimization and standard operating procedure. Nat. Protocols 1, 1507–1516. (Amine-modified microspheres, PAO5N [Cat. 450, 451]).
Lugli E, Gattinoni L, Roberto A, Mavilio D, Price DA, Restifo NR, Roederer M. (2012) Identification, isolation and in vitro expansion of human and nonhuman primate T stem cell memory cells. Nature Protocol v8(1). (SMPLX R-NH2 Beads [Cat. 450, 451], stained for LIVE/DEAD Aqua Comp Beads)
Estes, M. L., Mund, J. A., Ingram, D. A., & Case, J. (2010). Identification of endothelial cells and progenitor cell subsets in human peripheral blood. Current protocols in cytometry, 9-33.(Cat. 450, 451 Viability Dye Compensation Standard [Amine Polymer Microspheres, PA04N] [Cat. 450, 451]).

FITC / PE COMPENSATION STANDARD

The FITC / PE Compensation Standard includes four microsphere populations that are ~7-9µm in diameter: an Autofluor™ population, and single populations surface-labeled with FITC, PE, and FITC / PE. The Autofluor™ population is dyed with a low level of fluorophore to approximate the autofluorescence of an unstained resting lymphocyte. This kit is useful for establishing quadrant boundaries and compensation settings for analyses using FITC and PE fluorochromes.

Catalog Number	Description
820	FITC / PE Compensation Standard (Autofluor™, FITC-PE, FITC/PE)

FLUORESCENCE REFERENCE

Single-color Fluorescence Reference Standards for flow cytometry are labeled with specific fluorochromes to exhibit the same spectral characteristics as labeled cells. They may be used to QC a specific path of the optical system, to optimize filter sets for fluorophores, to establish test-specific Target Channel values for instrument set-up, and as general compensation or intensity standards.

Catalog Number	Description
890	Certified Blank™
897	Acridine Orange
886	Alexa Fluor® 488
887	Alexa Fluor® 647
901	Allophycocyanine (APC)
898	Chlorophyll
895	Cy™5
906	DAPI
913	Far-Out Red
891	Fluorescein (FITC),
894	Hoechst 33342
916	Pacific Blue™
908	PE-Cy™5
889	PE-Cy™7
909	PE-TR
892	Propidium Iodide
899	R-Phycoerythrin (PE)
905	T.M. Rhodamine
893	Texas Red® (TR)
915	Violet Laser

Fluorescence Quantitation

Our Quantum™ MESF and Quantum™ Simply Cellular^® beads are external standards that enable the standardization of fluorescence intensity units irrespective of cytometer and software. Because they are labeled with the same fluorochromes used to label cells, they provide a synchronous response to the environment. To use, beads are run on the same day and at the same PMT & compensation settings as samples to establish a calibration curve that relates instrument channel values and standardized fluorescence intensity units (MESF or ABC). Cell samples are then read against the curve for determination of expression (i.e. quantitation of the fluor signal from each cell population).

QUANTUM™ MESF

Our Quantum™ MESF kits contain one blank population and a series of either four fluorescent microsphere populations labeled with varying amounts of Alexa Fluor^® 488, Alexa Fluor^® 647, APC, PE-Cy™5, or R-PE, or five fluorescent microsphere populations labeled with varying amounts of FITC. The assignment of fluorescence intensity in Molecules of Soluble Fluorochrome (MESF) units is performed through direct comparison of fluorescence measurements from solutions of the pure fluorochrome with those from microspheres surface-labeled with the same fluorochrome. See PDS 849 to determine effective FP ratio. A free QuickCal ^® analysis template is provided with each kit to aid in determining the expression level of cells, and for evaluating instrument linearity and detection threshold. Quantum™ MESF kits are available in three sizes: A (20 tests), B (100 tests), and C (280 tests).

Notice to FITC-5 Customers: In our continual efforts to manufacture the most stable FITC MESF kits, we are utilizing an enhanced calibration strategy that features additional data points to best stabilize assignments through time. Though we anticipate that changes will not be evident for most customers, some may observe a slight shift in the calibration curve, observable in low or high assignments.

Catalog Number	Description
488	Quantum™ Alexa Fluor® 488 MESF
555	Quantum™ FITC-5 MESF
555p	Quantum™ FITC-5 MESF (Premix)
827	Quantum™ R-PE MESF
828	Quantum™ PE-Cy™5 MESF
647	Quantum™ Alexa Fluor® 647 MESF
823	Quantum™ APC MESF

REFERENCES

Wyant T, Lackey A, Green M. (2008) Validation of a flow cytometry based chemokine internalization assay for use in evaluating the pharmacodynamic response to a receptor agonist. J Transl Med; 6:76.
Quadrini KJ, Hegelund AC, Cortes KE, Xue C, Kennelly SM, Ji H, Högerkorp C-M, Mc Closky TW. (2016) Validation of a flow cytometry-based assay to assess C5aR receptor occupancy on neutrophils and monocytes for use in drug development. Cytometry B Clin Cytom; 90B:177-190.
Bray RA. (2013) Lymphocyte crossmatching by flow cytometry. Methods Mol Biol; 1034:285-96.
Smith ML, Chyla B, McKeegan E, Tahir SK (2016) Development of a flow cytometric method for quantification of BCL-2 family members in chronic lymphocytic leukemia and correlation with sensitivity to BCL-2 family inhibitors. Cytometry B Clin Cytom
Kay S, Herishanu Y, Pick M, Rogowski, O, Baron S, Naparstek E, Polliack A, Deutsch, V. R. (2006) Quantitative flow cytometry of ZAP-70 levels in chronic lymphocytic leukemia using molecules of equivalent soluble fluorochrome. Cytometry, 70B: 218–226.
Muller L, Mitsuhashi M, Simms P, Gooding WE, Whiteside TL (2016) Tumor-derived exosomes regulate expression of immune function-related genes in human T cell subsets. Sci Rep 6:20254.
Singh H, Figliola MJ, Dawson MJ, Olivares S, Zhang L, et al. (2013) Manufacture of Clinical-Grade CD19-Specific T Cells Stably Expressing Chimeric Antigen Receptor Using Sleeping Beauty System and Artificial Antigen Presenting Cells.PLoS ONE; 8(5):e64138
Homann S, Hofmann C, Gorin AM, Nguyen HCX, Huynh D, Hamid P, … & Roy SS. (2017) A novel rapid and reproducible flow cytometric method for optimization of transfection efficiency in cells. PloS one, 12(9), e0182941. (Quantum MESF to confirm and quantitate transfection efficiency featuring a fluorescently-labeled vector)

QUANTUM™ SIMPLY CELLULAR^®

Quantum™ Simply Cellular^® (QSC) kits are comprised of five microsphere populations that are ~7-9µm in diameter: one blank and four labeled with increasing amounts of antibody. Each coated population binds a specific number of monoclonal antibodies of the noted species, which is equal to its Antibody Binding Capacity (ABC) value. The investigator labels QSC beads using the same antibodies used to stain cells. A free QuickCal^® analysis template is provided with each kit to aid in determining the expression level of cells, and for evaluating instrument linearity and detection threshold. Quantum™ Simply Cellular^® kits are available in three sizes: A (20 tests), B (100 tests), and C (280 tests). For additional information about working with these products, see our product data sheet on Antibody Titration or check out PDS 818, Tips and Troubleshooting.

Catalog Number	Description
815	Quantum™ Simply Cellular® anti-Mouse IgG
816	Quantum™ Simply Cellular® anti-Human IgG
817	Quantum™ Simply Cellular® anti-Rat IgG

REFERENCES

Bringeland, G. H., Bader, L., Blaser, N., Budzinski, L., Schulz, A. R., Mei, H. E., … & Gavasso, S. (2019). Optimization of receptor occupancy assays in mass cytometry: Standardization across channels with QSC beads. Cytometry Part A, 95(3), 314-322.
Randlev B, Huang L-C, Watatsu M, Marcus M, Lin A, Shih S-J. (2010) Validation of a quantitative flow cytometer assay for monitoring HER-2/neu expression level in cell-based cancer immunotherapy products. Biologicals; 38(2):249-259.
McBean RS, Wilson B, Liew Y-W, Hyland CA, Flower RL. (2015) Quantitation of Lan antigen in Lan+, Lan+(w) and Lan- phenotypes. Blood Transfus; 13:662-5
Phillips A, Boghaert E, Vaidya K, Mitten M, Norvell S, Falls H, …Reilly A (2016) ABT-414, an Antibody–Drug Conjugate Targeting a Tumor-Selective EGFR Epitope Mol Cancer Ther 15(4)661-669
Tillotson B, Cho YK, Shusta E, (2013) Cells and Cell Lysates: A Direct Approach for Engineering Antibodies Against Membrane Proteins Using Yeast Surface Display Methods 60(1)27–37

SIMPLY CELLULAR^®

Our Simply Cellular^® standard consists of a single population of antibody-coated microspheres (~7-9µm in diameter) of known Antibody Binding Capacity (ABC). The population is stained with the user’s primary antibody and used in conjunction with a suitable Quantum™ MESF kit for determination of the antibody’s effective Fluorophore : Protein (F/P) ratio. This standard may also be used to QC the fluorescence intensity of different antibody lots or clones. Simply Cellular^® kits are available in three sizes: A (20 tests), B (100 tests), and C (280 tests). For additional information about working with these products, see our product data sheet on Antibody Titration or check out PDS 818, Tips and Troubleshooting.

Catalog Number	Description
810	Simply Cellular® anti-Mouse IgG
812	Simply Cellular® anti-Human IgG
813	Simply Cellular® anti-Rat IgG

REFERENCES

Andreev J, Thambi N, Perez Bay AE, Delfino F, Martin J, Kelly M… Thurston G. (2017) Bispecific Antibodies and Antibody–Drug Conjugates (ADCs) Bridging HER2 and Prolactin Receptor Improve Efficacy of HER2 ADCs. Mol Cancer Ther. (16)(4) 681-693 (Quantum MESF PE [quantitation] & Simply Cellular [F:P ratio])

QUICKCAL^®

QuickCal^® constructs a calibration curve associating the cytometer’s fluorescence channel values to standardized fluorescence intensity units. (MESF or ABC) For instructions regarding QuickCal, please see Product Data Sheet 819, QuickCal^® v. 2.3 Data Analysis Program or watch our video.

Cell Cycle Analysis

Bangs Laboratories offers a range of microspheres that may be used as standards to support assays in cell cycle analysis. Beads may be used for routine instrument set-up and QC, or as test-specific standards. Standards are available for use with flow cytometers, fluorescence microscopes, and cell viability analyzers. Custom products or preparations can also be manufactured to meet the requirements of specific assays or analyzers. For additional information regarding cell cycle analysis, see our Cell Cycle Analysis brochure.

FLUORESCENCE REFERENCE STANDARDS

Catalog Number	Description
897	Acridine Orange
906	DAPI
891	Fluorescein (FITC)
894	Hoechst 33342
892	Propidium Iodide

VIABILITY DYE COMPENSATION STANDARD

Viability Dye Compensation Standards are suitable for labeling with LIVE / DEAD^® stains or similarly-reactive dyes to generate compensation standards for flow cytometric analyses. Beads are not suitable for labeling with DNA stains such as propidium iodide, DAPI, or SYTOX^®, and users should contact us for discussion if uncertain as to the compatibility of a specific dye or stain.

Catalog Number	Description
450	Viability Dye Compensation Standard, 4µm
451	Viability Dye Compensation Standard, 8µm

Size Estimation

SMALL BEAD CALIBRATION KITS

Interest in exosome and microvesicle analysis has grown rapidly in recent years as another way to explore disease states, cancer progression, and for drug discovery and delivery. Due to their small size (30 – 100nm) however, these particles are difficult to detect using traditional methods, and require extra processing to segregate or characterize from debris commonly found in biological samples. Flow cytometry has emerged as a popular method of analysis and sorting for small particle characterization.

Catalog Number	Description
834	Nanobead Calibration Kit – 50nm, 100nm
832	Submicron Bead Calibration Kit – 0.2µm, 0.5µm, 0.8µm
833	Micron Bead Calibration Kit – 1.0µm, 3.0µm, 6.0µm

REFERENCES

Kong, F, Zhang, L, Wang, H, Yuan, G, Guo, A, Li, Q, & Chen, Z. (2015). Impact of collection, isolation and storage methodology of circulating microvesicles on flow cytometric analysis. Experimental and Therapeutic Medicine, 10, 2093-2101.
Huang S, Hou HW, Kanias T, et al. Towards microfluidic-based depletion of stiff and fragile human red cells that accumulate during blood storage. Lab on a chip. 2015;15(2):448-458.
Østergaard E, Frandsen PL, Sandberg E, (2015) Determination of freeze damage on HPV vaccines by use of flow cytometry. Biologicals Vol. 43,4:266–273
Venable AS, Williams RR, Haviland DL, McFarlin BK (2014) An analysis of endothelial microparticles as a function of cell surface antibodies and centrifugation techniques. J Immunol Methods, Vol406,117-123
Fischer S, Cornils K, Speiseder T, Badbaran A, Reimer R, Indenbirken, D, … & Lange, C. (2016). Indication of horizontal DNA gene transfer by extracellular vesicles. PLoS One, 11(9), e0163665.(Cat. 832, 833, Submicron & Micron Bead Calibration Kits; Cat. CC03N, SureCount 3µm)

Our Size Calibration Standards Kit for flow cytometry is comprised of five populations of undyed microspheres in the range of ~4-11µm. Forward scatter channel (FSC) values may be plotted against reported bead diameter to generate a curve that may be used for estimation of the sizes of cells run at the same instrument settings. The Size Calibration Standards Kits are available in two sizes: B (100 tests) and C (280 tests).

Catalog Number	Description
829	Size Calibration Standards Kit

REFERENCES

Mathew G, Timm Jr. EA, Sotomayor P, Godoy A, Montecinos VP, Smith G, Huss WJ (2009) ABCG2-mediated DyeCycle Violet efflux defined side population in benign and malignant prostate. Cell Cycle, 8:7, 1053-1061
Obokata H, Kojima K, Yamato M, Okano T, Tsuneda S, Vacanti C, (2011) The Potential of Stem Cells in Adult Tissues Representative of the Three Germ Layers Tissue Eng Part A 17(5-6):607-15

SIZE CALIBRATION STANDARDS

Cell Counting

Though the application of particles and cell counting technology is diverse, there is a common need for particulate standards. We manufacture count standards for both dedicated particle counters and flow cytometers. Enumeration of cells in a sample may be achieved through use of a microsphere count standard. Cell count may be calculated by determining the ratio of fluorescent count beads to unlabeled cells. For information on proper mixing and sampling, see TSD 0706: Handling & Pipetting Concentration Standards.

FLOW CYTOMETRY ABSOLUTE COUNT STANDARD™

Our Flow Cytometry Absolute Count Standard™ is a precisely counted population of microspheres for estimating counts of unlabeled cells via flow cytometry. The microspheres are ~7-9µm in diameter and supplied at ~1e+⁶beads/mL. Beads are internally labeled with multiple fluorophores for excitation with common lasers (e.g. 488nm, 633nm) and discrimination from the cell population. By evaluating the ratio of microspheres to cells, the volumetric number of cells may be determined.

Catalog Number	Nominal Mean Diameter (µm)
580	Flow Cytometry Absolute Count Standard™

REFERENCES

Døsen G, Tenstad E, Nygren MK, Stubberud H, Funderud S, Rian E. (2006) Wnt expression and canonical Wnt signaling in human bone marrow B lymphopoiesis. BMC Immunol. Jun 29;7:13.
Xiong, Z, Oriss, TB, Cavaretta, JP, Rosengart, MR, Lee, JS (2012) Red cell microparticle enumeration: validation of a flow cytometric approach. Vox Sang 103(1)p42–48 (Absolute Count standard; use with our Submicron Kit, Cat. 832)
Borzova, E, Dahinden, C, (2014) The Absolute Basophil Count. Meth Mol Biol V.1192 p87-100

Suspension Arrays

Suspension arrays for the flow cytometer feature populations of microspheres coated with different ligands for the interrogation of multiple targets within a given sample. Ligand-specific microsphere populations are encoded so that results for corresponding individual targets may be discerned. Encoding may be in the form of microsphere size and/or fluorescence signature, as with our QuantumPlex™ and QuantumPlex™M platforms.

Bangs’ QuantumPlex™ platform is an innovative bead platform specifically designed for suspension array applications. The beads may be analyzed using any standard flow cytometer (488nm or 633nm excitation), and do not require the use of specialized software.

QUANTUMPLEX™

Our QuantumPlex™ kits provide a general platform for multiplexed assay development on standard flow cytometers. Microsphere populations in 5 – bead kits are encoded with different intensities of Starfire Red™ and the two size kits (4.4 μm, 5.5 μm) may be combined to extend the array. Starfire Red™ emits in the red channel (e.g., PE-Cy™5, APC) with very little carry-over into lower wavelengths, leaving other detectors available for determination of positive binding events via common reporters such as FITC and PE. QuantumPlex™ kits are available with two different surfaces to accommodate the coating strategy of choice: carboxyl (COOH) and streptavidin (SA). Carboxyl populations are at a concentration of ~1×10⁸ beads/mL, while Streptavidin are at a concentration of ~1×10⁶ beads/mL. QuantumPlex™ are available in three sizes: A (1mL), B (5mL), and C (10mL).

Catalog Number	Description
235	QuantumPlex™ COOH 4.4 µm, 5 populations
238	QuantumPlex™ COOH 5.5 µm, 5 populations
239	QuantumPlex™ COOH 4.4 µm & 5.5 µm, 10 populations
215	QuantumPlex™ Streptavidin 4.4 µm, 5 populations
218	QuantumPlex™ Streptavidin 5.5 µm, 5 populations
219	QuantumPlex™ Streptavidin 4.4 µm & 5.5 µm, 10 populations

REFERENCES

Moss DM, Montgomery JM, Newland SV, Priest JW, Lammie PJ. (2004) Detection of cryptosporidium antibodies in sera and oral fluids using multiplex bead assay. J Parasitol; 90(2):397-404. [QuantumPlex COOH]
Yu J, Carvalho Mda G, Beall B, Nahm MH. (2008) A rapid pneumococcal serotyping system based on monoclonal antibodies and PCR. J Med Microbiol; 57(Pt 2):171-8. [QuantumPlex COOH]
Kellar, KL, Iannone, M (2002) Multiplexed microsphere-based flow cytometric assays. Exp. Hematol , V30, 11,p 1227-1237
Hale M, Lee B, Honaker Y, Leung W, Grier A, Jacobs H…Rawlings D. (2017) Homology-Directed Recombination for Enhanced Engineering of Chimeric Antigen Receptor T Cells. Mol Ther Methods Clin Dev; Vol 4 p192-203
Kellar KL, Mahmutovic AJ, Bandyopadhyay K. (2006) Multiplexed microsphere-based flow cytometric immunoassays. Curr Protoc Cytom. Feb;Chapter 13:Unit13.1.
Dolgushin SA, Odintsova ES, Gerasimenko AY, Tronin AV, Tereshchenko SA.(2015) Preliminary Tests of Multiplex Immunoassay for Detection of TORCH Infections in Human Blood Serum Using Flow Cytometry. Biomed Eng. Vol 49, 2,85-89
Spiro A, Lowe M, Brown D.(2000)A Bead-Based Method for Multiplexed Identification and Quantitation of DNA Sequences Using Flow Cytometry. Appl. Environ. Microbiol. Vol 66(10)p4258–4265

QUANTUMPLEX™ SP

QuantumPlex™ SP (Single Population) is useful for the development of simplex flow cytometric assays, or for the optimization of attachment chemistry and assay parameters before transitioning to a multiplexed format. Like QuantumPlex™, they are dyed with Starfire Red™ and are suitable for use on standard flow cytometers. Carboxyl (COOH) and streptavidin surfaces are available. Carboxyl populations are at a concentration of ~1×10⁸ beads/mL, while Streptavidin populations are at a concentration of ~1×10⁶ beads/mL. QuantumPlex™ SP are available in two sizes: A (1mL) or B (3mL).

Catalog Number	Description
234	QuantumPlex™ SP COOH 4.4 µm
237	QuantumPlex™ SP COOH 5.5 µm
214	QuantumPlex™ SP Streptavidin 4.4µm
217	QuantumPlex™ SP Streptavidin 5.5 µm

QUANTUMPLEX™M

QuantumPlex™M (Magnetic) kits consist of five populations of highly uniform ~6µm superparamagnetic microspheres encoded with different intensities of Starfire Red™. Like QuantumPlex™, they are suitable for use on standard flow cytometers. Highly efficient separations may be performed using rare earth magnetic separators. Carboxyl (COOH) and streptavidin surfaces are available. Carboxyl populations are at a concentration of ~1×10⁸ beads/mL, while Streptavidin populations are at a concentration of ~1×10⁶ beads/mL. QuantumPlex™M are available in three sizes: A (1mL), B (5mL), and C (10mL).

Catalog Number	Description
250	QuantumPlex™M COOH, 5 populations
252	QuantumPlex™M Streptavidin, 5 populations

REFERENCES

Yang S, Lien K, Huang K, Lei H, Lee G. (2008) Micro flow cytometry utilizing a magnetic bead-based immunoassay for rapid virus detection. Biosens. Bioelectron Vol 24:4 855-862 (QuantumPlex M Streptavidin)
Joslin J, Gilligan J, Anderson P, Garcia C, Sharif O, Hampton J … Lesley S (2018) A Fully Automated High-Throughput Flow Cytometry Screening System Enabling Phenotypic Drug Discovery. 05/29 SLAS DISC: Adv Life Sci R&D (QuantumPlex M COOH)

QUANTUMPLEX™M SP

QuantumPlex™M SP (Magnetic, Single Population) is useful for the development of simplex flow cytometric assays, or for the optimization of attachment chemistry and assay parameters before transitioning to a multiplexed format. Like QuantumPlex™M, they are highly uniform ~6µm superparamagnetic microspheres dyed with Starfire Red™ and are suitable for use on standard flow cytometers. Carboxyl (COOH) and streptavidin surfaces are available. Carboxyl populations are at a concentration of ~1×10⁸ beads/mL, while Streptavidin are at a concentration of ~1×10⁶ beads/mL. QuantumPlex™M SP are available in two sizes: A (1mL) or B (3mL).

Catalog Number	Description
251	QuantumPlex™M SP COOH
253	QuantumPlex™M SP Streptavidin

Fluorescence Reference

FLUORESCENCE REFERENCE

Single-color Fluorescence Reference Standards for flow cytometry are labeled with the same common fluorophores that are used to label cells. They may be used to QC a specific path of the optical system, to optimize filter sets for specific tests (e.g. immunophenotyping, cell viability, cell cycle, etc), and as general compensation or intensity standards. See our flow cytometry fluorophore reference chart.

Catalog Number	Description
890	Certified Blank™
897	Acridine Orange
886	Alexa Fluor® 488
887	Alexa Fluor® 647
901	Allophycocyanine (APC)
898	Chlorophyll
895	Cy™5
906	DAPI
913	Far-Out Red
891	Fluorescein (FITC),
894	Hoechst 33342
916	Pacific Blue™
908	PE-Cy™5
889	PE-Cy™7
909	PE-TR
892	Propidium Iodide
899	R-Phycoerythrin (PE)
905	T.M. Rhodamine
893	Texas Red® (TR)
915	Violet Laser

For additional trademark information, see our Trademarks page.

REFERENCES

Cretich, M., Di Carlo, G., Giudici, C., Pokoj, S., Lauer, I., Scheurer, S., & Chiari, M. (2009). Detection of allergen specific immunoglobulins by microarrays coupled to microfluidics. Proteomics, 9(8), 2098-2107.(Propidium Iodide FRS)
Franklin, RB, Campbell, AH, Higgins, CB, Barker, MK, & Brown, BL (2011). Enumerating bacterial cells on bioadhesive coated slides. Journal of microbiological methods, 87(2), 154-160.(Cat. 892, Propidium Iodide FRS)

Flow Cytometry

QUANTUM™ QC, 8 PEAK BEADS

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SMALL BEAD CALIBRATION KITS

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FLOW CYTOMETRY ABSOLUTE COUNT STANDARD™

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QUANTUMPLEX™

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QUANTUMPLEX™ SP

QUANTUMPLEX™M

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QUANTUMPLEX™M SP

FLUORESCENCE REFERENCE

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