PE Mouse Anti-Human LAP

BD Pharmingen™ PE Mouse Anti-Human LAP

Name: PE Mouse Anti-Human LAP
Brand: PE Mouse Anti-Human LAP
SKU: 562260

Clone TW4-2F8 (RUO)

Multicolor flow cytometric analysis of LAP expression on human peripheral blood lymphocytes. Human peripheral blood mononuclear cells were activated (24 h) with plate-bound Purified NA/LE Mouse Anti-Human CD3 (Cat. No. 555329) and Purified NA/LE Mouse Anti-Human CD28 (Cat. No. 555725) antibodies. The cells were harvested and stained with PerCP Mouse Anti-Human CD4 (Cat. No. 347324) and either PE Mouse IgG1, κ Isotype Control (Cat. No. 555749; Left Panel) or PE Mouse Anti-Human LAP (Cat. No. 562260; Right Panel) antibodies. The cells were then fixed and stained with the Alexa Fluor® 488 Mouse Anti-Human FoxP3 (Cat. No. 560047) antibody according to the recommended protocol. Two-color flow cytometric dot plots showing the correlated expression patterns of FoxP3 versus LAP (or Ig isotype control staining) were derived from CD4 positive-gated cells with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System. Intracellular staining for LAP on P3UI-TGFβ1 transfectant has also been demonstrated.

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Product Details

Brand: BD Pharmingen™

Alternative Name: Latency-associated peptide; TGFB1; TGFbeta; Transforming growth factor beta

Reactivity: Human (QC Testing)

Isotype: Mouse BALB/c IgG1, κ

Immunogen: Human TGF-β1

Application: Intracellular staining (flow cytometry) (Routinely Tested), Flow cytometry (Tested During Development)

Vol. Per Test: 5 µl

RRID: AB_10895568

Storage Buffer: Aqueous buffered solution containing BSA and ≤0.09% sodium azide.

Regulatory Status: RUO

Preparation And Storage

Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. 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.

Recommended Assay Procedures

Suggested Staining Procedures for PE Mouse anti-Human LAP Antibody:

1. Harvest the PBMCs after stimulation (24 hours) with plate-bound Purified NA/LE Mouse Anti-Human CD3 (Cat. No. 555329) and

Purified NA/LE Mouse Anti-Human CD28 (Cat. No. 555725).

2. Wash the cells twice with stain buffer (eg. BD Pharmingen™ Stain Buffer (FBS), Cat. No. 554656).

3. Stain 1 × 10^6 cells with PerCP Mouse anti-Human CD4 (Cat. No. 347324) and either with the PE Mouse anti-Human LAP antibody

(Cat. No. 562260) or with PE Mouse IgG1, κ Isotype control (Cat. No. 555749) for 30 minutes on ice, protected from light.

4. Wash cells twice with stain buffer.

5. Stain for Alexa Fluor® 488 Mouse anti-Human FoxP3, refer to Technical Data Sheet of Cat. No. 560047 for detailed protocol.

In brief,

a. Add 2 ml of 1 × FoxP3 buffer A to the cell pellet.

b. Centrifuge and incubate in 0.5 ml of buffer C for 30 minutes.

c. Wash cells twice with stain buffer and stain with anti-FoxP3 antibody for 30-45min.

d. Wash cells twice with stain buffer and acquire on the Flow cyotmeter.

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).
An isotype control should be used at the same concentration as the antibody of interest.
For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
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.
Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.

Companion Products

Stain Buffer (FBS) RUO

Size 500 mL Cat No. 554656

Lysing Buffer RUO

Size 100 mL Cat No. 555899

Human FoxP3 Buffer Set RUO

Size 100 Tests Cat No. 560098

PE Mouse IgG1, κ Isotype Control RUO

Size 100 Tests Cat No. 555749

Purified NA/LE Mouse Anti-Human CD3 RUO

Size 0.5 mg Cat No. 555329

Purified NA/LE Mouse Anti-Human CD28 RUO

Size 0.5 mg Cat No. 555725

View Details

562260 Rev. 3

Antibody Details

TW4-2F8

The TW4-2F8 monoclonal antibody specifically binds to Latency-Associated Peptide (LAP), a component of the dimeric Transforming Growth Factor-beta 1 (TGF-β1) propeptide encoded by TGFB1. Prior to secretion, the dimeric LAP-TGF-β1 propeptide is cleaved resulting in a biologically inactive form of dimeric TGF-β1 that is noncovalently associated with dimeric LAP (latent TGF-β1). This complex may be expressed on the surface of TGF-β1-producing cells or be further processed by proteolytic removal of LAP to release the biologically active mature form of the soluble TGF-β1 homodimer. Platelets contain TGF-β1 and most nucleated cells, including tumor cells and cells that comprise the innate and adaptive immune system can produce TGF-β1. TGF-β1 is a potent multifunctional cytokine that regulates numerous processes including development, hematopoiesis, tissue remodeling, wound repair, and immunity as well as cancer and autoimmune diseases. Clone TW4-2F8 is routinely quality tested through intracellular staining of LAP in P3UI-TGFβ1 transfected cells.

562260 Rev. 3

Format Details

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.

Format: PE

Excitation Source: Yellow-Green 488 nm, 532 nm, 561 nm

Excitation Max: 496 nm, 566 nm

Emission Max: 576 nm

562260 Rev.3

Citations & References

View product citations for antibody "562260" on CiteAb

Development References (2)

Oida T, Weiner HL. Overexpression of TGF-β1 gene induces cell surface localized glucose-regulated protein 78-associated latency-associated peptide/TGF-β. J Immunol. 2010; 185(6):3529-3535. (Clone-specific: Flow cytometry, Immunoprecipitation, Western blot). View Reference
Rubtsov YP, Rudensky AY. TGFbeta signalling in control of T-cell-mediated self-reactivity. Nat Rev Immunol. 2007; 7(6):443-453. (Biology). View Reference

562260 Rev. 3

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.