Multicolor flow cytometric analyses of XBP-1S expressed in human and mouse cells. Panel 1. Analysis of XBP-1S expressed in activated human peripheral blood mononuclear cells (PBMC). Human PBMC were stimulated with 10 μg/ml ODN 2006 Type B CpG Oligonucleotide (Invivogen, Cat. No. tlrl-2006-1) at 37˚C for 7 days. The cells were harvested and incubated with BD Horizon™ Fixable Viability Stain 450 (Cat. No. 562247), fixed in 1X Fix/Perm Buffer (BD Pharmingen™ Transcription Buffer Set, Cat: 562574) at 2-8˚C (45 min) and permeabilized in 1X Perm/Wash Buffer (Transcription Buffer Set) at 2-8˚C (45 min). Cells were stained with PE Mouse anti-XBP-1S (Cat. No. 562642) and PerCP-Cy™5.5 Mouse Anti-Human CD20 (Cat. No. 558021) antibodies. Two-color flow cytometric contour plots showing the correlated expression of CD20 versus XBP-1S (or Ig isotype control staining) were derived from live-gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry was performed using a BD FACSCanto™ II Flow Cytometer System. Panel 2. Analysis of XBP-1S expressed in activated mouse splenocytes. BALB/C splenocytes were stimulated with 1 μg/ml lipopolysaccharide (LPS; Sigma Cat. No. L3137) at 37˚C (3 days). Cells were incubated with Fixable Viability Stain 450, fixed in 1X Fix/Perm Buffer, and permeabilized in 1X Perm/Wash Buffer as described for Panel 1. The cells were stained with PE Mouse anti-XBP-1S and APC Rat Anti-Mouse CD45R/B220 (BD Cat. No. 553092). The cells were then analyzed as described above. Contour plots showing the correlated expression of B220 versus XBP-1S (or Ig isotype control staining) were derived from live-gated events with the forward and side light-scatter characteristics of intact lymphocytes.
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Multicolor flow cytometric analyses of XBP-1S expressed in human and mouse cells. Panel 1. Analysis of XBP-1S expressed in activated human peripheral blood mononuclear cells (PBMC). Human PBMC were stimulated with 10 μg/ml ODN 2006 Type B CpG Oligonucleotide (Invivogen, Cat. No. tlrl-2006-1) at 37˚C for 7 days. The cells were harvested and incubated with BD Horizon™ Fixable Viability Stain 450 (Cat. No. 562247), fixed in 1X Fix/Perm Buffer (BD Pharmingen™ Transcription Buffer Set, Cat: 562574) at 2-8˚C (45 min) and permeabilized in 1X Perm/Wash Buffer (Transcription Buffer Set) at 2-8˚C (45 min). Cells were stained with PE Mouse anti-XBP-1S (Cat. No. 562642) and PerCP-Cy™5.5 Mouse Anti-Human CD20 (Cat. No. 558021) antibodies. Two-color flow cytometric contour plots showing the correlated expression of CD20 versus XBP-1S (or Ig isotype control staining) were derived from live-gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry was performed using a BD FACSCanto™ II Flow Cytometer System. Panel 2. Analysis of XBP-1S expressed in activated mouse splenocytes. BALB/C splenocytes were stimulated with 1 μg/ml lipopolysaccharide (LPS; Sigma Cat. No. L3137) at 37˚C (3 days). Cells were incubated with Fixable Viability Stain 450, fixed in 1X Fix/Perm Buffer, and permeabilized in 1X Perm/Wash Buffer as described for Panel 1. The cells were stained with PE Mouse anti-XBP-1S and APC Rat Anti-Mouse CD45R/B220 (BD Cat. No. 553092). The cells were then analyzed as described above. Contour plots showing the correlated expression of B220 versus XBP-1S (or Ig isotype control staining) were derived from live-gated events with the forward and side light-scatter characteristics of intact lymphocytes.
Product Details
BD Pharmingen™
XBP1; X-box binding protein 1; TREB5; Tax-responsive element-binding
Mouse (QC Testing), Human (Tested in Development)
Mouse IgG1, κ
Human XBP-1S Recombinant Protein
Intracellular staining (flow cytometry) (Routinely Tested)
5 µl
AB_2687882
Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
RUO
Preparation And Storage
The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated with R-PE under optimum conditions, and unconjugated antibody and free PE were removed. Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.
Recommended Assay Procedures
Note: For optimal immunofluorescent staining of XBP1-S, the Transcription Factor Buffer Set (Cat. No. 562574) is highly recommended for the brightest staining results.
Product Notices
- This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
- Source of all serum proteins is from USDA inspected abattoirs located in the United States.
Companion Products
Stain Buffer (FBS) RUO
Size
500 mL
Cat No.
554656
Fixable Viability Stain 450 RUO
Size
0.1 mg
Cat No.
562247
Transcription Factor Buffer Set RUO
Size
100 Tests
Cat No.
562574
PE Mouse IgG1, κ Isotype Control RUO
Size
0.1 mg
Cat No.
554680
PerCP-Cy™5.5 Mouse Anti-Human CD20 RUO
Size
50 Tests
Cat No.
558021
APC Rat Anti-Mouse CD45R/B220 RUO
Size
0.1 mg
Cat No.
553092
562642 Rev. 1
Antibody Details
Q3-695
The Q3-695 monoclonal antibody specifically binds to XBP-1S, the spliced isoform of X-box binding protein 1 (XBP-1). XBP-1S is generated by cells that undergo an unfolded protein response, ie, cells responding to the accumulation of unfolded proteins in the endoplasmic reticulum. It plays important roles as a transcription factor in a number of important cellular processes including the regulation of MHC class II genes, differentiation of plasma cells, immunoglobulin secretion and hepatocyte growth.
562642 Rev. 1
Format Details
PE
R-Phycoerythrin (PE), is part of the BD family of Phycobiliprotein dyes. This fluorochrome is a multimeric fluorescent phycobiliprotein with excitation maximum (Ex Max) of 496 nm and 566 nm and an emission maximum (Em Max) at 576 nm. PE is designed to be excited by the Blue (488 nm), Green (532 nm) and Yellow-Green (561 nm) lasers and detected using an optical filter centered near 575 nm (e.g., a 575/26-nm bandpass filter). As PE is excited by multiple lasers, this can result in cross-laser excitation and fluorescence spillover on instruments with various combinations of Blue, Green, and Yellow-Green lasers. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
PE
Yellow-Green 488 nm, 532 nm, 561 nm
496 nm, 566 nm
576 nm
562642 Rev.1
Citations & References
Development References (3)
-
Gass JN, Gifford NM, Brewer JW. Activation of an unfolded protein response during differentiation of antibody-secreting B cells. J Biol Chem. 2002; 277(50):49047-49054. (Biology). View Reference
-
Iwakoshi NN, Lee AH, Vallabhajosyula P, Otipoby KL, Rajewsky K, Glimcher LH. Plasma cell differentiation and the unfolded protein response intersect at the transcription factor XBP-1. Nat Immunol. 2003; 4(4):321-329. (Biology). View Reference
-
Yoshida H, Matsui T, Yamamoto A, Okada T, Mori K. XBP1 mRNA is induced by ATF6 and spliced by IRE1 in response to ER stress to produce a highly active transcription factor. Cell. 2001; 107(7):881-891. (Biology). View Reference
562642 Rev. 1
Please refer to Support Documents for Quality Certificates
Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described
Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims. Comparisons are not made against non-BD technologies, unless otherwise noted.
For Research Use Only. Not for use in diagnostic or therapeutic procedures.
