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AdipoGen Life Sciences
Calreticulin (human) (rec.) (His)
230
CHF
CHF 230.00
In stock
AG-40A-0132-C01010 µgCHF 230.00
AG-40A-0132-C05050 µgCHF 590.00
Figure 1: Flow cytometric analysis of CRT on the cell surface
3.105 EL4 Thymoma cells, growing in suspension in RPMI 1640 (Gibco) supplemented medium were plated in 12-well plates and treated with mitomycin C (30mM, Sanofi Aventis) or cisplatin (25mM, Sigma) for 4h. Cells were harvested, washed once with cold PBS and possibly resuspended in 200mL of cold PBS containing 1mg of recombinant Calreticulin (Prod. No. AG-40A-0132, Adipogen) for 30 minutes on ice. After one wash with cold PBS, cells were fixed in 0.25% paraformaldehyde (PFA) in PBS for 5 minutes. After washing again once with cold PBS, cells were incubated for 30min with primary antibody, diluted in cold blocking buffer (2% FBS in PBS), followed by washing and incubation with the Alexa488-conjugated monoclonal secondary antibody in blocking buffer (30 min). Each sample was then analyzed by FACScan (Becton Dickinson) to identify cell-surface Calreticulin. Secondary antibody alone was used as an isotype control, and the fluorescent intensity of stained cells was gated on propidium iodide (PI) negative cells.Pictures courtesy of Prof. Guido Kroemer, INSERM, Paris.
Figure 2:
Immunofluorescence
Cells were possibly incubated with rCRT (Prod. No. AG-40A-0132, Adipogen) and stained with the same protocol than for FACS analysis. Then, cells resuspended in 100mL of PBS were seeded on polylysine slides for 15 min then fixed with 100mL of 4% PFA add on the cells for 15 min. Drops were gently aspirated before using the mounting medium including DAPI from Vectashield. Slides were then analyzed by confocal microscopy. Oxaliplatin (150mM, Sanofi Aventis) and mitoxanthron (1mM, Sigma) treated cells were used as positive control. Pictures courtesy of Prof. Guido Kroemer, INSERM, Paris.
| Product Details | |
|---|---|
| Synonyms | CRP55; Calregulin; Endoplasmic Reticulum Resident Protein 60; ERp60; grp60 |
| Product Type | Protein |
| Properties | |
| Source/Host | E. coli |
| Sequence |
Human calreticulin (aa18-417) is fused at the C-terminus to a His-tag. |
| Crossreactivity | Human |
| MW | ~55kDa (SDS-PAGE) |
| Purity | ≥90% (SDS-PAGE) |
| Purity Detail | Purified using Ni-NTA column and FPLC. |
| Endotoxin Content | <1EU/μg purified protein (LAL test; Lonza). |
| Concentration | 0.2mg/ml |
| Formulation | Liquid. 0.2μm-filtered solution in 55mM TRIS-Cl, pH 8.2, containing 150mM NaCl. |
| Other Product Data |
UniProt link P27797: Calreticulin (human) |
| Shipping and Handling | |
| Shipping | BLUE ICE |
| Short Term Storage | +4°C |
| Long Term Storage | -20°C |
| Handling Advice |
After opening, prepare aliquots and store at -20°C. Avoid freeze/thaw cycles. For maximum product recovery after thawing, centrifuge the vial before opening the cap. |
| Use/Stability |
Stable for at least 6 months after receipt when stored at -20°C. Working aliquots are stable for up to 3 months when stored at -20°C. |
| Documents | |
| MSDS |
 Download PDF |
| Product Specification Sheet | |
| Datasheet |
Download PDF |
Description
Calreticulin is involved in regulation of intracellular Ca2+ homoeostasis and ER Ca2+ capacity. It constitutes together with calnexin and ERp57 the 'calreticulin/calnexin cycle' that is responsible for folding and quality control of newly synthesized glycoproteins. Calreticulin has been implicated to play a role in many biological systems, including functions inside and outside the ER, indicating that the protein is a multi-process molecule. Calreticulin was shown to enhance the merger of macrophages and tumor cells, increasing phagocytosis. Recently, calreticulin has been identified as an endogenous ligand of the NK cell receptor NKp46. NKp46 recognizes externalized calreticulin (ecto-CRT), which translocates from the endoplasmic reticulum (ER) to the cell membrane during ER stress. ER stress and ecto-CRT are hallmarks of chemotherapy-induced immunogenic cell death, favivirus infection and senescence. NKp46-mediated killing is inhibited by knockout or knockdown of CALR, the gene encoding CRT, or CRT antibodies, and is enhanced by ectopic expression of glycosylphosphatidylinositol-anchored CRT. NKp46 recognition of ecto-CRT as a danger-associated molecular pattern eliminates ER-stressed cells.