Detection of Human GILT/IFI30 by Western Blot.
Western blot shows lysates of HepG2 human hepatocellular carcinoma cell line. PVDF membrane was probed with 0.5 µg/mL of Sheep Anti-Human GILT/IFI30 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7715) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for GILT/IFI30 at approximately 25-30 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.
GILT/IFI30 in Human Spleen.
GILT/IFI30 was detected in immersion fixed paraffin-embedded sections of human spleen using Sheep Anti-Human GILT/IFI30 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7715) at 1 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Sheep HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS019) and counterstained with hematoxylin (blue). Specific staining was localized to lysosomes in cytoplasm. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.
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.
IFI30 (Gamma-interferon-inducible protein IP-30; also gamma-interferon [IFN-gamma ] inducible lysosomal thiol reductase/GILT and Legumaturain) is a 25-30 kDa member of the GILT family of proteins. It is constitutively expressed in B cells and dendritic cells, and induced by IFN-gamma in non-APCs. IFI30 is both intracellular and secreted as an inactive glycosylated proenzyme. The glycosylation pattern contains a terminal phosphorylated mannose, which is recognized by cell surface mannose-6 phosphate receptors and internalized into lysosomes. In lysosomes, IFI30 is processed into an active, mature form, and via a thiol reductase domain, breaks disulfide bonds in molecules destined for lysosomal degradation. This is a critical first step in the processing and subsequent presentation of peptides that will initiate an antigenic response. The human IFI30 proenzyme is 224 amino acids (aa) in length. In this form, it is 33-35 kDa in size. Following proteolytic processing at both the N- and C-terminus, a 175 aa, 25-30 kDa active mature form is generated (aa 58-232). The mature region possesses a thiol reductase domain (aa 62-151) plus one utilized Thr phosphorylation site. Both the pro- and mature forms exhibit enzymatic activity. IFI30 is known to exist as a 50-60 kDa disulfide-linked homodimer. There are four potential isoform variants. One contains a 26 aa substitution for aa 213-250, a second shows a deletion of aa 131-161, a third shows a deletion of aa 106-123, while a fourth shows a deletion of aa 64-212. Over aa 27-250, human IFI30 shares 62% aa sequence identity with mouse IFI30.
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