Anti-GRIM19 antibody [6E1BH7]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-GRIM19 antibody [6E1BH7] (AB110240)

Mitochondrial localization of GRIM19 visualized by immunocytochemistry using ab110240 at a concentration of 1 µg/mL. Cultured Human fibroblasts were fixed, permeabilized and then labeled with ab110240 followed by Alexa® 488 goat-anti-mouse IgG.

• Flow Cyt

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Flow Cytometry - Anti-GRIM19 antibody [6E1BH7] (AB110240)

HeLa cells were stained with 1 µg/mL ab110240 (blue) or an equal amount of an isotype control antibody (red) and analyzed by flow cytometry.

• WB

Lab

Western blot - Anti-GRIM19 antibody [6E1BH7] (AB110240)

Lanes 1-3 : Merged signal (red and green). Green - ab110240 observed at 17 kDa. Red - loading control ab181602 observed at 36 kDa.

ab110240 GRIM19 antibody [6E1BH7] was shown to specifically react with GRIM19 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265863 (knockout cell lysate ab257136) was used. Wild-type and GRIM19 knockout samples were subjected to SDS-PAGE. ab110240 and Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 680RD) preadsorbed (ab216777) and Goat anti-Mouse IgG H&L (IRDye^® 800CW) preadsorbed (ab216772) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-GRIM19 antibody [6E1BH7] (ab110240) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

NDUFA13 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human NDUFA13 (GRIM19) knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-ndufa13-grim19-knockout-hela-cell-line-ab265863'>ab265863</a>)

Lane 3:

Jurkat cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-680rd-preadsorbed-ab216777'>ab216777</a>) at 1/10000 dilution

Predicted band size: 17 kDa,184 kDa,28 kDa,56 kDa,62 kDa,76 kDa

Observed band size: 17 kDa,184 kDa,29 kDa,56 kDa,70 kDa,75 kDa

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• WB

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Western blot - Anti-GRIM19 antibody [6E1BH7] (AB110240)

All lanes:

Western blot - Anti-GRIM19 antibody [6E1BH7] (ab110240) at 1 µg/mL

Lane 1:

Human heart mitochondria

Lane 2:

Bovine heart mitochondria

Lane 3:

Rat heart mitochondria

Lane 4:

Mouse heart mitochondria

Predicted band size: 17 kDa

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• WB

CiteAb

Western blot - Anti-GRIM19 antibody [6E1BH7] (AB110240)

Western Blotting using Anti-GRIM19 antibody [6E1BH7], ab110240. Publication image from Kim, M. J. et al., 2023, Nat Commun, 37433777. Legend direct from paper.

Small HSP and HSP110 families have a tendency to be increased under mitochondrial stress.a RNA-seq analysis of HSPs gene expression log2 fold changes (log2FC) in NDUFA11 KO and NDUFA13 KO compared to WT HEK293T cells (n = 4). Up- and down-regulated genes (q-value < 0.05) are shown in green and pink, respectively. The intensity of the color shades depends on the level of expression change. Gray indicates genes with not statistically significant expression changes. b mRNA expression patterns of selected transcripts validated by RT-qPCR. The mRNA levels are presented as fold changes relative to WT. Data shown are mean ± SD (n = 3 biological replicates with two technical replicates). p-value from an ordinary one-way ANOVA with Dunnett’s multiple comparisons test using GraphPad Prism. c Western blot analysis of HSPs expression performed in whole cell lysates of NDUFA11 KO, NDUFA13 KO and WT HEK293T cells. ACTB was used as a loading control. Data shown are representative of three independent experiments. d Quantification of HSPs in western blot analysis normalized to ACTB using ImageJ. The protein levels are presented as fold changes relative to WT. Data shown are mean ± SD (n = 3). p-value from two-sided, unpaired t-test using GraphPad Prism. Source data are provided as a Source Data file.

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• WB

CiteAb

Western blot - Anti-GRIM19 antibody [6E1BH7] (AB110240)

Western Blotting using Anti-GRIM19 antibody [6E1BH7], ab110240. Publication image from Kopajtich, R. et al., 2017, Nat Commun, 28604674. Legend direct from paper.

RNA aberrant expression detection and validation.(a) Aberrantly expressed genes (Hochberg corrected P value<0.05 and |Z-score|>3) for each patient fibroblasts. (b) Gene-wise RNA expression volcano plot of nominal P values (−log10P value) against Z-scores of the patient #35791 compared against all other fibroblasts. Z-scores with absolute value >5 are plotted at ±5, respectively. (c) Same as (b) for patient #73804. (d) Sample-wise RNA expression is ranked for the genes TIMMDC1 (top) and MGST1 (bottom). Samples with aberrant expression for the corresponding gene are highlighted in red (#35791, #66744, and #73804). (e) Gene-wise comparison of RNA and protein fold changes of patient #35791 compared to the average across the fibroblast cell lines of all other patients. Subunits of the mitochondrial respiratory chain complex I are highlighted (red squares). Reliably detected proteins that were not detected in this sample are shown separately with their corresponding RNA fold changes (points below solid horizontal line). (f) Western blot of TIMMDC1, NDUFA13, NDUFB3 and NDUFB8 protein in three fibroblast cell lines without (#62346, #91324, NHDF) and three with a variant in TIMMDC1 (#35791, #66744 and #96687), and fibroblasts re-expressing TIMMDC1 (‘-T’) (#35791-T, #66744-T and #96687-T). UQCRC2 was used as loading control. CI, complex I subunit; CIII, complex III subunit; MW, molecular weight. (g) Blue native PAGE blot of the control fibroblasts re-expressing TIMMDC1 (NHDF-T), the control fibroblasts (NHDF), patient fibroblasts (#96687) and patient fibroblast re-expressing TIMMDC1 (#96687-T). Immunodecoration for complex I and complex III was performed using NDUFB8 and UQCRC2 antibodies, respectively. CI, complex I subunit; CIII, complex III subunit.

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