Grp78

Manufactured by Enzo Life Sciences

Sourced in United States

GRP78 is a molecular chaperone protein that plays a critical role in the endoplasmic reticulum (ER) stress response. It is involved in the folding and assembly of proteins within the ER lumen, as well as the regulation of the unfolded protein response (UPR).

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GRP78/BiP ELISA kit (2 citations) GRP78)/BiP (2 citations)

6 protocols using grp78

Biomarker Quantification Protocol

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FETUB (abcam^®, Cambridge, MA, USA) and GRP-78 (BioCell PC, Enzo Life Sciences AG, Switzerland) were measured by ELISA in all subjects using a 96-well plate immunoassay kit according to the manufacturers’ protocol. Plasma fibrinogen concentration was measured by the Clauss method29 (link). Concentrations of ALT and AST were measured by a biochemistry automatic analyser (Roche Diagnostics, Branchburg, NJ, USA).

Diao W.Q., Shen N., Du Y.P., Liu B.B., Sun X.Y., Xu M, & He B. (2016). Fetuin-B (FETUB): a Plasma Biomarker Candidate Related to the Severity of Lung Function in COPD. Scientific Reports, 6, 30045.

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Multiparameter Serum Biomarker Assays

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ELISA assays for IGFBP2 and LCAT (Cloud Clone Corp., Wuhan, Hubei, China), SHBG (R&D Systems, Abingdon, UK), GRP78 (Enzo Life Sciences, Exeter, UK) and calprotectin (Biolegend, San Diego, CA, USA) were performed in duplicate using commercial kits following manufacturers’ instructions.
The mean coefficients of variance for duplicate analysis for each assay were as follows: IGFBP2: 8.1% LCAT: 8.4% SHBG: 7.4% GRP78: 3.1% and calprotectin: 4.5%. Serum CA125 levels were available as previously described (Menon et al, 2009a (link)).

Russell M.R., Graham C., D'Amato A., Gentry-Maharaj A., Ryan A., Kalsi J.K., Ainley C., Whetton A.D., Menon U., Jacobs I, & Graham R.L. (2017). A combined biomarker panel shows improved sensitivity for the early detection of ovarian cancer allowing the identification of the most aggressive type II tumours. British Journal of Cancer, 117(5), 666-674.

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Multimarker Immunofluorescence Staining Protocol

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Primary antibodies used for immunostaining included those for EGFP (Santa Cruz Biotechnology), MMP7 (R & D Systems, Minneapolis, MN, USA), cytokeratin (Leica, Solms, Germany), CD34 (Abcam, Cambridge, MA, USA), GRP78 (Enzo Life Sciences, Lausen, Switzerland), cleaved caspase-3, cleaved caspase-4, cleaved caspase-9, cleaved caspase-12, Cox IV (all from Cell Signaling Technology, Danvers, MA, USA); secondary antibodies included goat anti-mouse HRP (Santa Cruz Biotechnology), goat anti-rabbit HRP (Santa Cruz Biotechnology), donkey anti-goat HRP (Santa Cruz Biotechnology), chicken anti-rat AP (Santa Cruz Biotechnology), chicken anti-rabbit AP (Santa Cruz Biotechnology), chicken anti-mouse AP (Santa Cruz Biotechnology), goat anti-rabbit Alexa 488/594 (Invitrogen), rabbit anti-goat Alexa 594 (Invitrogen), chicken anti-rabbit Alexa 594 (Invitrogen) and chicken anti-mouse Alexa 594 (Invitrogen). Immunostaining was performed using these antibodies. The sections were counter-stained with hematoxylin or nuclear fast red (Vector, Burlingame, CA, USA). For double staining, EGFP, MMP7, cytokeratin, CD34 and PAS staining was performed following TUNEL staining or cleaved caspase-3 staining.

Zhan Y., Xu C., Liu Z., Yang Y., Tan S., Yang Y., Jiang J., Liu H., Chen J, & Wu B. (2016). β-Arrestin1 inhibits chemotherapy-induced intestinal stem cell apoptosis and mucositis. Cell Death & Disease, 7(5), e2229-.

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Quantifying Serum GRP78 Levels in Clinical Samples

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Blood samples were collected during patients’ routine outpatient visit. Samples were kept on ice and centrifugated at 1500 × g for 15 min. Subsequently, serum was transferred into coded aliquots and stored at −80°C. Analysis of GRP78 levels (Enzo Life Sciences, Inc, Farmingdale, NY, USA) was conducted by commercially available enzyme-linked immunosorbent assay (ELISA) kits. 4-Parameters Logistic Regression (Graphpad Prism 9.0.0) was applied to analyse GRP concentrations.
In addition to GRP measurement, blood samples were acquired to assess full blood count and high-sensitivity C-reactive protein (hs-CRP) levels.

Al Zaidi M., Marggraf V., Repges E., Nickenig G., Skowasch D., Aksoy A, & Pizarro C. (2023). Relevance of serum levels of the endoplasmic reticulum stress protein GRP78 (glucose-regulated protein 78 kDa) as biomarker in pulmonary diseases. Cell Stress & Chaperones, 28(3), 333-341.

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Biomarker Profiling in COVID-THELIUM

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The present study is an ancillary study of the COVID-THELIUM cohort. Assays were performed using an Enzyme-Linked Immunosorbent Assay for sRAGE, VEGF-A (Invitrogen, ThermoFisher Scientific, Waltham, MA, USA) and GRP78 (Enzo Life Sciences, Villeurbanne, France), and using an Electro-Chemiluminescence Immunoassay for IL-6 (Roche Diagnostics, Meylan, France).

Besnier E., Brunel V., Thill C., Leprêtre P., Bellien J., Demailly Z., Renet S., Tamion F, & Clavier T. (2022). Soluble RAGE as a Prognostic Marker of Worsening in Patients Admitted to the ICU for COVID-19 Pneumonia: A Prospective Cohort Study. Journal of Clinical Medicine, 11(15), 4571.

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Quantifying Plasma Protein Biomarkers in PAH

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Blood was drawn from a peripheral vein of PAH patients routinely presenting to our outpatient clinic and collected directly into EDTA tubes. The samples were kept on ice and centrifuged (1500×g, 15 min at 4°C) for a maximum of 30 min. Thereafter, plasma was transferred into coded aliquots and frozen at −80°C until the time of analysis.
The plasma concentrations of GRP78 (Enzo Life Sciences, Inc, Farmingdale, New York, USA) and HSP27 (R&D Systems, Bio-Techne, Minneapolis, USA) were analysed by using commercially available enzyme-linked immunosorbent assay (ELISA) kits. 4-Parameters Logistic Regression (Graphpad Prism 9.0.0) was used to plot the concentrations.

Al Zaidi M., Pizarro C., Bley C., Repges E., Sedaghat A., Zimmer S., Jansen F., Tiyerili V., Nickenig G., Skowasch D, & Aksoy A. (2022). ER-stress-induced secretion of circulating glucose-regulated protein 78kDa (GRP78) ameliorates pulmonary artery smooth muscle cell remodelling. Cell Stress & Chaperones, 27(5), 561-572.

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