Human TGF-beta 2 DuoSet ELISA

R&D Systems | Catalog # DY302

R&D Systems
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Key Product Details

Assay Type

Solid Phase Sandwich ELISA

Assay Range

31.2-2000 pg/mL

Sample Type

Cell culture supernates, serum, and plasma
Note: Diluents for complex matrices, such as serum and plasma, should be evaluated prior to use in this DuoSet

Reactivity

Human

Human TGF-beta 2 DuoSet ELISA Features

  • Optimized capture and detection antibody pairings with recommended concentrations save lengthy development time
  • Development protocols are provided to guide further assay optimization
  • Assay can be customized to your specific needs
  • Economical alternative to complete kits
View all TGF-beta 2 ELISA Kits »

Product Summary for Human TGF-beta 2 DuoSet ELISA

This DuoSet ELISA Development kit contains the basic components required for the development of sandwich ELISAs to measure natural and recombinant human TGF-beta2. The suggested diluent is suitable for the analysis of most cell culture supernate samples. Diluents for complex matrices, such as serum and plasma, should be evaluated prior to use in this DuoSet.

 

Product Specifications

Assay Format

96-well strip plate (sold separately)

Sample Volume Required

100 µL

Detection Method

Colorimetric ELISA - 450nm (TMB)

Conjugate

Biotin

Specificity

Please see the product datasheet

Label

HRP

Scientific Data Images for Human TGF-beta 2 DuoSet ELISA

Human TGF-beta 2 ELISA Standard Curve

Human TGF-beta 2 ELISA Standard Curve

Kit Contents for Human TGF-beta 2 DuoSet ELISA

  • Capture Antibody
  • Detection Antibody
  • Recombinant Standard
  • Streptavidin conjugated to horseradish-peroxidase (Streptavidin-HRP)

Other Reagents Required

DuoSet Ancillary Reagent Kit 2 (5 plates): (Catalog # DY008C) containing 96 well microplates, plate sealers, substrate solution, stop solution, plate coating buffer (PBS), wash buffer, and Reagent Diluent Concentrate 2.

PBS: (Catalog # DY006), or 137 mM NaCl, 2.7 mM KCl, 8.1 mM Na2HPO4, 1.5 mM KH2PO4, pH 7.2 - 7.4, 0.2 µm filtered

Wash Buffer: (Catalog # WA126), or equivalent

Reagent Diluent*

Blocking Buffer*

Substrate Solution: ELISA TMB Substrate (Catalog # DY999B or DY999B-250)

Stop Solution: Methanesulfonic acid (Catalog # DY994B or DY994B-250)

Microplates: (Catalog # DY990), or equivalent

Plate Sealers: (Catalog # DY992), or equivalent

*For the recommended Reagent Diluent and Blocking Buffer for a specific DuoSet ELISA Development Kit, refer to the product datasheet.

Preparation and Storage

Shipping

The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store the unopened product at 2 - 8 °C. Do not use past expiration date.

Background: TGF-beta 2

TGF-beta 2 is synthesized as a prepro-cytokine with a 19 amino acid (aa) signal sequence, a 283 aa pro-region, and a 112 aa mature segment (1-5). It dimerizes with formation of disulfide bonds between the 'pro' regions and disulfide bonds between the 'mature' regions. The mature region is 71% and 80% identical with human TGF-beta 1 and TGF-beta 3 (6, 7) and 97% identical with the corresponding mouse protein (8). After proteolytic cleavage of the disulfide-linked mature region, it remains hydrogen-bonded to the disulfide-linked prosegments (LAP or latencyassociated protein) (1, 2, 9). If secreted in this form, LAP keeps TGF-beta 2 in an inactive state until dissociation, caused by proteases, glycosidases, or extreme pH (2, 9). In many types of cells, an additional protein, latent TGF-beta binding protein (LTBP), is covalently linked to the LAP homodimer prior to secretion. LTBP, a 130 kDa cysteine-rich glycoprotein, creates a 235 kDa large latent complex that is secreted, most likely binding to the extracellular matrix (1, 9-11). The latency components are believed to act as natural antagonists of TGF-beta activity, to target TGF-beta to distinct tissues, and to maintain a reservoir of TGF-beta (1, 2, 12). On release from latency, active homodimeric TGF-beta can bind to cell-surface receptors or to other proteins, such as alpha 2-macroglobulin (13). 
The signal transducing receptor for TGF-beta 2 is a heterotetrameric complex of two type I signal-transducing receptors (53 kDa; TGF-beta RI) and two type II ligand-binding receptors (75-85 kDa; TGF-beta RII) (14-19). The binding of TGF-beta 2 appears to initially involve a type III TGF-beta receptor, either 300 kDa betaglycan (20) or 180 kDa endoglin (14, 21), which then "hands off" to TGF-beta RII. 
TGF-beta 2 is expressed by a variety of cells, including osteoclasts, thymic epithelium, keratinocytes, hepatocytes, chief cells of the stomach, satellite cells, skeletal muscle cells, prostatic epithelium, bronchial epithelium, neurons and astrocytes, fibroblasts and visceral smooth muscle, and macrophages (14-19). TGF-beta 2 has marked cross-species bioactivity (e.g., human TGF-beta 2 is active on mouse cells (33), while porcine TGF-beta 2 is active on rabbit cells (34)). TGF-beta 2 has four fundamental activities: it is a growth inhibitor for most types of cells; it enhances the deposition of extracellular matrix; it is immunosuppressive, suppressing APC expression of both IL-12 and CD40L while upregulating IL-10 secretion; and during fetal development, it is expressed in discrete areas, such as epithelium, myocardium, cartilage and bone of extremities and in the nervous system, suggesting specific functions (1, 35-37).

Long Name

Transforming Growth Factor beta 2

Alternate Names

TGFB2, TGFbeta 2

Entrez Gene IDs

7042 (Human); 21808 (Mouse); 397084 (Porcine)

Gene Symbol

TGFB2

Additional TGF-beta 2 Products

Product Documents for Human TGF-beta 2 DuoSet ELISA

Certificate of Analysis

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Note: Certificate of Analysis not available for kit components.

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Product Specific Notices for Human TGF-beta 2 DuoSet ELISA

For research use only

Citations for Human TGF-beta 2 DuoSet ELISA

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Protocols

View specific protocols for Human TGF-beta 2 DuoSet ELISA (DY302):

GENERAL ELISA PROTOCOL

Plate Preparation

  1. Dilute the Capture Antibody to the working concentration in PBS without carrier protein. Immediately coat a 96-well microplate with 100 μL per well of the diluted Capture Antibody. Seal the plate and incubate overnight at room temperature.
  2. Aspirate each well and wash with Wash Buffer, repeating the process two times for a total of three washes. Wash by filling each well with Wash Buffer (400 μL) using a squirt bottle, manifold dispenser, or autowasher. Complete removal of liquid at each step is essential for good performance. After the last wash, remove any remaining Wash Buffer by aspirating or by inverting the plate and blotting it against clean paper towels.
  3. Block plates by adding 300 μL Reagent Diluent to each well. Incubate at room temperature for a minimum of 1 hour.
  4. Repeat the aspiration/wash as in step 2. The plates are now ready for sample addition.

Assay Procedure

  1. Add 100 μL of sample or standards in Reagent Diluent, or an appropriate diluent, per well. Cover with an adhesive strip and incubate 2 hours at room temperature.
  2. Repeat the aspiration/wash as in step 2 of Plate Preparation.
  3. Add 100 μL of the Detection Antibody, diluted in Reagent Diluent, to each well. Cover with a new adhesive strip and incubate 2 hours at room temperature.
  4. Repeat the aspiration/wash as in step 2 of Plate Preparation.
  5. Add 100 μL of the working dilution of Streptavidin-HRP to each well. Cover the plate and incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
  6. Repeat the aspiration/wash as in step 2.
  7. Add 100 μL of Substrate Solution to each well. Incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
  8. Add 50 μL of Stop Solution to each well. Gently tap the plate to ensure thorough mixing.
  9. Determine the optical density of each well immediately, using a microplate reader set to 450 nm. If wavelength correction is available, set to 540 nm or 570 nm. If wavelength correction is not available, subtract readings at 540 nm or 570 nm from the readings at 450 nm. This subtraction will correct for optical imperfections in the plate. Readings made directly at 450 nm without correction may be higher and less accurate.

 

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