Detection of VISTA/B7-H5/PD-1H by Western Blot.
Western blot shows lysates of mouse platelets. PVDF membrane was probed with 1 µg/mL of Sheep Anti-Mouse VISTA/B7-H5/PD-1H Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7005) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for VISTA/B7-H5/PD-1H at approximately 30 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 8.
Detection of VISTA/B7-H5/PD-1H in HEK293 Human Cell Line Transfected with Mouse VISTA and eGFP by Flow Cytometry.
HEK293 human embryonic kidney cell line transfected with either (A) mouse VISTA or (B) irrelevant transfectants and eGFP was stained with Sheep Anti-Mouse VISTA/B7-H5/PD-1H Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7005) followed by Allophycocyanin-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # F0127). Quadrant markers were set based on control antibody staining (Catalog # 5-001-A). View our protocol for Staining Membrane-associated Proteins.
VISTA/B7-H5/PD-1H in D3 Mouse Cell Line.
VISTA/B7-H5/PD-1H was detected in immersion fixed D3 mouse embryonic stem cell line using Sheep Anti‑Mouse VISTA/B7-H5/PD-1H Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7005) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Sheep IgG Secondary Antibody (red; Catalog # NL010) and counterstained with DAPI (blue). Specific staining was localized to cell surfaces and cytoplasm. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.
Preparation and Storage
Sterile PBS to a final concentration of 0.2 mg/mL.
Reconstitution Buffer Available
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below. *Small pack size (SP) is shipped with polar packs. Upon receipt, store it immediately at -20 to -70 °C
Stability & Storage
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
12 months from date of receipt, -20 to -70 °C as supplied.
1 month, 2 to 8 °C under sterile conditions after reconstitution.
6 months, -20 to -70 °C under sterile conditions after reconstitution.
Platelet Receptor Gi24 (also known as VISTA, B7-H5, SISP1, C10orf54 and, Dies1 [Differentiation of ESC-1]) is a 55-65 kDa member of the Ig superfamily. It is a transmembrane molecule expressed in bone, on embryonic stem cells (ESCs), and on tumor cell surfaces. On ESCs, Gi24 appears to positively interact with BMP4, potentiating BMP signaling and the transition from an undifferentiated to a differentiated state. On tumor cells, Gi24 both promotes MT1-MMP expression and activity, and serves as a substrate for MT1-MMP. This increases the potential for cell motility. Mature mouse Gi24 is a 277 amino acid (aa) type I transmembrane glycoprotein (aa 33-309). It contains a 149 aa extracellular region (aa 33-191) with one V-type Ig-like domain (aa 33-161) and a 97 aa cytoplasmic domain. Based on human Gi24, mouse Gi24 will likely undergo proteolytic cleavage by MT1-MMP, generating a soluble 30 kDa extracellular fragment, plus a 25-30 kDa membrane-bound fragment. There are two potential isoform variants. One contains a deletion of aa 127-187, while another shows an alternative start site at Met82. Over aa 33-191, mouse Gi24/Dies1 shares 78% and 70% aa identity with rat and human Gi24, respectively.
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