BD ACCURI NEWS

 

March 2017


Spotlight

New BD Accuri™ product manager Mara Swaim

Spotlight - Swaim Interview

Mara Swaim is the new product manager for the BD Accuri™ product line. She joined BD in 2013 after managing several labs at Johns Hopkins University. We caught up with Mara to talk about her background, her enthusiasm for flow cytometry and what she likes most about her new job.
Read the interview »

Noteworthy

The 2017 BD Horizon™ Global Tour is coming!

Noteworthy - Horizon Tour Thumb

Replay our BD Horizon webinars for essential information, expert tips and considerations for multicolor flow cytometry. You’ll learn how to Reveal, Elevate, Resolve, Achieve and use the Sentinel approach. Then stay tuned for the 2017 BD Horizon™ Global Tour, with dates and cities to be announced soon.
View the videos »

Application Highlight

Cytokine analysis using BD™ CBA

Cytokines and growth factors—one of the broad range of life science applications discussed in our new white paper–are a primary means of communication between cells, driving cellular differentiation that develops and supports the adaptive immune system. Different cytokines result in different cellular responses, which in turn cause production of more cytokines. Measurement of cytokines and growth factors can provide valuable information about immune responses and cell functionality. However, running multiple conventional ELISA assays of single analytes can consume substantial time, labor, budget and sample material.

Bead-based flow cytometric immunoassays are powerful methods for quantifying cytokines because they allow researchers to multiplex many analytes with very little sample. A BD™ Cytometric Bead Array (CBA) assay (Figure 1) starts with a cocktail of beads bound with specific capture antibodies that can be detected by a flow cytometer due to slight differences in fluorescence intensity. The beads are mixed with samples of plasma, serum or tissue culture supernatants, along with PE-labeled detection antibodies, forming a sandwich complex. The beads are washed to eliminate unbound proteins and then run on a BD Accuri C6 Plus or other flow cytometer. Finally, data files are exported to FCAP Array™ software for analysis.

App Highlight - Fig 1 Workflow Diagram

Figure 1. Overview of BD CBA workflow

BD CBA can analyze 300 beads per cytokine—the equivalent of 300 ELISA wells—for up to 30 cytokines simultaneously. In essence, BD CBA is like running multiple ELISAs at once using flow cytometry.

There are two ways to use BD CBA: kits and flex sets. BD CBA Kits provide preconfigured panels of 3–7 analytes for ultimate ease of use. For example, the BD™ CBA Human Th1/Th2/Th7 Cytokine Kit shown in Figure 2 can quantitate IL-2, IL-4, IL-6, IL-10, TNF, IFN-γ and IL-17A simultaneously with a two-color assay. Other kits include panels of human cytokines, chemokines and anaphylatoxins; mouse cytokines and immunoglobulins; and non-human primate cytokines.

App Highlight - Fig 2 Cytokine with CBA kits Thumb
Figure 2. Analysis of cytokine expression with BD CBA kits
App Highlight - Fig 2 Cytokine with CBA kits
Figure 2. Analysis of cytokine expression with BD CBA kits

BD CBA assays can quantify multiple cytokines simultaneously using minimal sample. In this experiment, seven cytokines were quantified in culture supernatant of stimulated peripheral blood mononuclear cells using the BD CBA Human Th1/Th2/Th17 Cytokine Kit (Cat. No. 560484). Capture beads for each cytokine were identified in FL4 of the BD Accuri C6 Plus, and cytokine levels were measured based on bead-signal intensities in FL2. Cytokine concentrations were calculated using standard curves.

On the BD Accuri C6 Plus, the beads are excited by the red laser and detected in FL4, while the PE reporter is excited by the blue laser and detected in FL2. A free downloadable BD Accuri™ C6 Plus software template simplifies setup and acquisition. Results can be displayed in FCAP Array software by bead (graphing one analyte across all samples) or by sample (graphing one sample across all analytes).1

BD CBA Flex Sets are three-color assays that enable analysis of up to 30 analytes. Available specificities include soluble protein assays for detection of human, mouse or rat cytokines, chemokines and growth factors; human immunoglobulins; cell signaling assays; and enhanced sensitivity assays that can detect protein concentrations of 1.0 pg/mL or lower. The beads contain two dyes excited by the red laser that locate them on a 2D matrix, while a PE reporter again quantifies the analytes. Flex sets are completely configurable, and the workflow essentially allows researchers to build their own multiplex kit.

App Highlight - Fig 3 Cytokine with CBA Flex Sets Thumb
Figure 3. Analysis of cytokine expression with BD CBA flex sets
App Highlight - Fig 3 cytokine with CBA Flex Sets
Figure 3. Analysis of cytokine expression with BD CBA flex sets

Human peripheral blood mononuclear cells were cultured for several days with plate-bound anti-CD3, soluble anti-CD27, IL-2 and IL-4. In one experimental condition, cells were stimulated with PMA+Ionomycin for several hours before harvesting. In the other, cells were stimulated for several hours with IFN-γ, and LPS was added to the culture overnight. All samples were prepared using the BD™ CBA Human Soluble Protein Master Buffer Kit (Cat. No. 558264) and stained with flex set reagents for ten different cytokines. Samples were acquired on a BD Accuri C6 in the 2-blue, 2-red configuration using the BD™ CBA Flex Set Accuri Template and analyzed using FCAP Array software v3.0.1. Results: The bar chart shows all ten cytokine levels for one sample under both experimental conditions.

Flex sets require the optional Selectable Laser module (Cat. No. 661085) to reconfigure the BD Accuri C6 Plus so that two detectors (instead of the usual one) read signal from the red laser. Again, a free downloadable software template simplifies setup and acquisition, and data analysis in FCAP Array is similar to kits. Figure 3 illustrates how you could compare production of ten different cytokines by blood cells in response to two different stimulation regimes.

BD CBA assays assess the total concentration of a given cytokine without identifying which subset of cells secreted it. An alternative approach, intracellular cytokine staining (ICS), enables researchers to interrogate distinct subpopulations of a heterogeneous sample at the single-cell level for expression of a cytokine of interest. You’ll find examples of ICS in our product information sheet on analyzing cytokine expression in cancer cell lines. Together, these two complementary approaches can provide a more comprehensive understanding of cytokine kinetics.

Download the new BD Accuri life sciences white paper »

Download our product information sheet on intracellular cytokine staining »

Learn more about BD CBA »


 

Tips & Tricks

Resources for BD Accuri C6 customers

Tips and Tricks - Accuri C6 Plus

With last year’s introduction of the next-generation BD Accuri C6 Plus personal flow cytometer, sales of BD Accuri™ C6 instruments have been discontinued globally. BD is absolutely committed to supporting your needs by providing best-in-class service and support to BD Accuri™ products. Current warranties are not affected and will be supported until they expire. Service and support, as well as product options and accessories, are available for purchase and provided by BD as usual and without compromise.

You will find a full range of documentation, product information sheets, technical documents, webinars and downloadable software templates on the BD Accuri C6 Resources page on bdbiosciences.com. To access the information, click BD Accuri C6 System on the right panel of any BD Accuri C6 Plus page—or just click the link below.

Visit the BD Accuri C6 Resources page »


 

Publication Picks

This section highlights interesting recent research articles using BD Accuri flow cytometers.

New cancer drugs

Sharma P, Srinivasa Reddy T, Thummuri D, et al. Synthesis and biological evaluation of new benzimidazole-thiazolidinedione hybrids as potential cytotoxic and apoptosis inducing agents. Eur J Med Chem. 2016;124:608-621. PubMed

Immune changes after typhus infection

Papp S, Moderzynski K, Rauch J, et al. Liver necrosis and lethal systemic inflammation in a murine model of Rickettsia typhi infection: role of neutrophils, macrophages and NK cells. PLoS Negl Trop Dis. 2016;10:e0004935. PubMed

Measurement of DNA damage

Ma L, Qiao Y, Jones R, Singh N, Su M. Single cell HaloChip assay on paper for point-of-care diagnosis. Anal Bioanal Chem. 2016;408:7753-7759. PubMed

Nanofiber production

Sun X, Li K, Chen S, et al. Rationally designed particle preloading method to improve protein delivery performance of electrospun polyester nanofibers. Int J Pharm. 2016;512:204-212. PubMed


 

Events

Meeting – April 1–5, 2017 – Washington, DC

AACR 2017 (American Association for Cancer Research) »

Meeting – May 12–16, 2017 – Washington, DC

Immunology 2017 (American Association of Immunologists) ›


Class 1 Laser Product.
For Research Use Only. Not for use in diagnostic or therapeutic procedures.

1 All reagents and kits are compatible with both the BD Accuri C6 and the new BD Accuri C6 Plus flow cytometer systems. Data was generated on the BD Accuri C6 Plus except where noted. Information about BD reagent kits, BD Accuri™ C6 and BD Accuri C6 Plus software templates and BD CSampler™ and BD CSampler Plus automation options is available at www.bdbiosciences.com/accuri.