Catalog Number: AF3080
Lot Number: WJV01
Size: 100 μg
Formulation: 0.2 μm filtered solution in PBS with 5% trehalose
Storage: -20° C
Reconstitution: sterile PBS
Specificity: human A33 extracellular domain
Immunogen: NS0-derived rhA33 extracellular domain
Ig Type: goat IgG
Applications: Western blot Immunohistochemistry
HT-29 cells were stained with anti-A33 (R&D Systems, Cat. # AF3080, filled histogram) or control antibody (R&D Systems, Cat. # AB-108-C, open histogram) followed by PE-conjugated anti-goat antibody (R&D Systems, Cat. # F0107).
Produced in goats immunized with purified, NS0-derived, recombinant human A33 (rhA33; aa 22 - 235; Accession # Q99795). Human A33 specific IgG was purified by human A33 affinity chromatography. Human A33, also known as GPA33, is a 43 kDa type I transmembrane glycoprotein that belongs to the CTX (cortical thymocyte marker in Xenopus) family of cell adhesion molecules within the immunoglobulin superfamily. Other family members include CXADR, ESAM, BT-IgSF, CD2 and JAM A-C. The extracellular domain (ECD) of human A33 is 214 amino acids (aa) in length and contains one V-type and one C2-type Ig-like domain. This ECD is 80%, 74% and 71% aa identical to canine, bovine and mouse A33 ECD, respectively. A33 is likely to be involved in cell-cell adhesion between epithelial cells.
Lyophilized from a 0.2 μm filtered solution in phosphate-buffered saline (PBS) with 5% trehalose.
Reconstitute with sterile PBS. If 0.5 mL of PBS is used, the antibody concentration will be 0.2 mg/mL.
Lyophilized samples are stable for twelve months from date of receipt when stored at -20° C to -70° C. Upon reconstitution, the antibody can be stored at 2° - 8° C for 1 month without detectable loss of activity. Reconstituted antibody can also be aliquotted and stored frozen at -20° C to -70° C in a manual defrost freezer for six months without detectable loss of activity. Avoid repeated freeze-thaw cycles.
This antibody has been selected for its ability to recognize human A33 in the applications listed below. In direct ELISAs and Western blots, this antibody shows approximately 10% cross-reactivity with rmA33.
Western blot - This antibody can be used at 0.1 - 0.2 μg/mL with the appropriate secondary reagents to detect human A33. The detection limit for rhA33 is approximately 2 ng/lane under non-reducing and reducing conditions.
Immunohistochemistry - This antibody will detect A33 in cells and tissues. The working dilution is 2 - 15 μg/mL. For chromogenic detection of labeling, use R&D Systems’ Cell and Tissue Staining Kits (CTS Series).
Flow Cytometry - This antibody was tested in flow cytometry using HT-29 cells. Dilute this antibody to 50 μg/mL and add 10 μL of this diluted solution to 1-2.5 x 105 cells in a total reaction volume not exceeding 200 μL. The binding of unlabeled antibodies may be visualized by adding a secondary developing reagent such as anti-goat IgG conjugated to a fluorochrome.
Direct ELISA - This antibody can be used at 0.5 - 1.0 μg/mL with the appropriate secondary reagents to detect human A33. The detection limit for rhA33 is approximately 0.2 ng/well.
Optimal dilutions should be determined by each laboratory for each application.