- PA5-19503 - Invitrogen Antibodies HSPA5 antibody

Submitted references Enhancing calmodulin binding to ryanodine receptor is crucial to limit neuronal cell loss in Alzheimer disease.

Nakamura Y, Yamamoto T, Xu X, Kobayashi S, Tanaka S, Tamitani M, Saito T, Saido TC, Yano M
Scientific reports 2021 Mar 31;11(1):7289

Effect of vitamin B(12) supplementation on retinal lesions in diabetic rats.

Reddy SS, Prabhakar YK, Kumar CU, Reddy PY, Reddy GB
Molecular vision 2020;26:311-325

Dantrolene prevents hepatic steatosis by reducing cytoplasmic Ca(2+) level and ER stress.

Tamitani M, Yamamoto T, Yamamoto N, Fujisawa K, Tanaka S, Nakamura Y, Uchinoumi H, Oda T, Okuda S, Takami T, Kobayashi S, Sakaida I, Yano M
Biochemistry and biophysics reports 2020 Sep;23:100787

Flutamide Alters the Expression of Chemerin, Apelin, and Vaspin and Their Respective Receptors in the Testes of Adult Rats.

Brzoskwinia M, Pardyak L, Rak A, Kaminska A, Hejmej A, Marek S, Kotula-Balak M, Bilinska B
International journal of molecular sciences 2020 Jun 22;21(12)

The role of vaspin in porcine corpus luteum.

Kurowska P, Mlyczyńska E, Dawid M, Grzesiak M, Dupont J, Rak A
The Journal of endocrinology 2020 Dec;247(3):283-294

Expression and Impact of Vaspin on In Vitro Oocyte Maturation through MAP3/1 and PRKAA1 Signalling Pathways.

Kurowska P, Mlyczyńska E, Estienne A, Barbe A, Rajska I, Soból K, Poniedziałek-Kempny K, Dupont J, Rak A
International journal of molecular sciences 2020 Dec 8;21(24)

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of CHO-K1 Whole Cell Lysate using Product # PA5-19503, GRP78 BiP primary antibody at a dilution of 1 µg/mL (lane 1). Staining of HeLa Whole Cell Lysate at a dilution of 1 µg/mL (lane 2). Blot treated with a secondary HRP-conjugated Goat polyclonal anti-Rabbit antibody was used at a dilution of 1:3000.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using Anti-GRP78 Polyclonal Antibody (Product # PA5-19503) and a 72 kDa band corresponding to GRP78 was observed across cell lines and tissue extracts tested and increased upon Thapsigargin treatment in Hela and NIH/3T3. Whole cell extracts (30 µg lysate) of HeLa (Lane 1), HeLa treated with Thapsigargin (1uM for 24hr) (Lane 2), NIH/3T3 (Lane 3) and NIH/3T3 treated with Thapsigargin (1uM for 24hr) (Lane 4) and tissue extracts of Mouse Liver (Lane 5), Rat Liver (Lane 6) and Mouse Pancreas (Lane 7) were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0322BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1 ug/ml) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L), Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005)..

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical (formalin-fixed, paraffin-embedded) staining of Human Liver Carcinoma tissue using Product # PA5-19503, anti-GRP78 BiP antibody. Primary antibody was used at a concentration of 1 µg/mL and exposed for 8 mins at room temp. The sample was pretreated using heat mediated antigen retrieval with Sodium Citrate Buffer (pH6/20mins). The detection method was a HRP conjugated polymer, DAB chromogen and the sample was counterstained with haematoxylin and mounted with DPX.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 1 Qualitative ( A - H , A' - H' ) and quantitative ( I ) analyses of immunohistochemical staining of chemerin ( A , A' ), CCRL2 ( B , B' ), CMKLR1 ( C , C' ), GPR1 ( D , D' ), apelin ( E , E' ), APLNR ( F , F' ), vaspin (G , G') and GRP78 (H , H') . Representative microphotographs of control (A - H) and flutamide-treated testes ( A' - H' ). Additionally stained with Mayer's haematoxylin. Bars = 10 mum. Positive staining for chemerin, CCRL2, CMKLR1, apelin and APLNR are restricted to Leydig cells (arrows), whereas staining of GPR1, vaspin and GRP78 is visible in Leydig cells (arrows) and seminiferous tubules (open arrows). Note the decreased levels of chemerin ( A' ), GPR1 ( D' ), APLNR ( F' ) and vaspin ( G' ) and increased levels of CCRL2 ( B' ) and apelin ( E' ) intensities after flutamide administration. The intensity of CMKLR1 and GRP78 staining in flutamide ( C',H' ) and control ( C , H ) Leydig cells are similar. No immunopositive staining of chemerin, apelin and vaspin and their receptors was observed when the primary antibodies are omitted ( A - H , A' - H', insertion in upper left corner of microphotographs). Histograms ( I ) depicting staining intensities of chemerin, apelin, vaspin, and their receptors are expressed as relative optical density of the brown staining product in Leydig cells. The results are presented as means +- SD. Statistically significant differences are flagged with asterisks (* p < 0.05, ** p < 0.01, *** p < 0.001). Control ( n = 6) and f

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 2 Representative Western blots assessing the relative expression of chemerin ( A ), CCRL2 ( B ), CMKLR1 ( C ), GPR1 ( D ) , apelin ( E ), APLNR (F), vaspin ( G ) and GRP78 ( H ) proteins in control and flutamide-treated testes. Normalisation was performed with beta-Actin as a loading control. Protein levels within control testes were given a value of 1. Data obtained from three separate analyses are presented as means +- SD. Statistically significant differences are flagged with asterisks (* p < 0.05, ** p < 0.01) between control ( n = 6) and flutamide-treated ( n = 6) animals; C: control, F: flutamide.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 2 78-kDa glucose-regulated protein (GRP78) mRNA and protein expression before and after oocyte in vitro maturation, as well as its immunolocalisation in cumulus-oocyte complexes (COCs). COCs (50/group/experiment) were selected after morphological examination of material collected from the ovaries taken from 100 pigs (15 follicles/ovary). COCs were collected before (0 h) and after (44 h) in vitro maturation and oocytes were denuded by gently pipetting in hyaluronidase, then real-time PCR and Western blot analysis were performed to determine mRNA ( A ) and protein ( B ) level of GRP78 in oocytes and cumulus cells. Immunolocalisation of GRP78 was studied by immunofluorescence in COCs ( C ). Gene expression level was normalised to the geometric mean of actin and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and protein to actin. GRP78 immunostaining shown by arrows. Experiments were performed independently and repeated three times ( n = 3). The data are plotted as the mean +- SEM of three independent experiments. Significance between groups before and after maturation is indicated by ** p < 0.01 and *** p < 0.001; Cumulus cells (Cc), oocyte (Oo), Immunoglobulin G (IgG).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 2 Protein expression of ER stress markers, TAU and p-TAU in WT, App NL-G-F , RYR2 V3599K , and App NL-G-F / RYR2 V3599K incubated neuronal cells. Representative images of ( A ) GRP78, ( B ) ATF6, ( C ) CHOP, ( D ) TAU/p-TAU obtained from single cells, and the summarized data. Merged images with DAPI staining are also shown. p-TAU was expressed as the ratio to total TAU. Cell shapes are indicated by dotted lines. Values for individual mice are plotted together with mean +- SEM. Scale bars: 30 mum. N = 20-33 cells from 3 to 6 mice. Parentheses indicate the number of mice. * p < 0.05, ** p < 0.01, *** p < 0.001 (one-way ANOVA with post-hoc Tukey's multiple comparison test).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 6 Immunoblotting for ER stress and apoptotic markers in the rat retina. A : Representative immunoblots for endoplasmic reticulum (ER) stress markers and apoptotic markers. B : Quantification of the corresponding densitometry data. The protein expression was normalized to the tubulin and was represented as %control. Data are mean +- standard error of the mean (SEM, n=3). C, control; D, diabetes; D+B12, diabetic rats treated with vitamin B 12 . *Significant difference from the control group; # significant difference from the diabetes group (p

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Fig. 4 GRP78 in hepatocytes. Immunocyte-fluorescent study of GRP78 was done for primary murine hepatocytes after incubation for 24 h in either media alone (CNT group), media containing 0.2 mM palmitate (FFA group), or media containing 0.2 mM palmitate and 3 muM dantrolene (FFA + DAN group). A: Representative images of immunocyte-fluorescent study for GRP78 hepatocytes. B: GRP78 fluorescent intensity were calculated. There were significant differences between FFA group and FFA + DAN group. Values are the mean +- SEM of 60 cells from three different experiments. Fig. 4

PA5-19503

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