Anti-SSRP1 抗体 [10D7]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-SSRP1 antibody [10D7] (AB26212)

ICC/IF image of ab26212 stained HepG2 cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab26212, 10µg/ml) overnight at +4°C. The secondary antibody (green) was ab96879, DyLight® 488 goat anti-mouse IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

• Flow Cyt

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Flow Cytometry - Anti-SSRP1 antibody [10D7] (AB26212)

Overlay histogram showing HeLa cells stained with ab26212 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab26212, 1μg/1x10⁶ cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG2b [PLPV219] (ab91366, 2μg/1x10⁶ cells ) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HeLa cells fixed with 4% paraformaldehyde (10 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.

• WB

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Western blot - Anti-SSRP1 antibody [10D7] (AB26212)

All lanes:

Western blot - Anti-SSRP1 antibody [10D7] (ab26212) at 1 µg/mL

All lanes:

HeLa cell lysate

Secondary

All lanes:

Goat anti-mouse Ig conjugated to HRP

Predicted band size: 81 kDa

Observed band size: 81 kDa

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• WB

CiteAb

Western blot - Anti-SSRP1 antibody [10D7] (AB26212)

SSRP1 western blot using anti-SSRP1 antibody [10D7] ab26212. Publication image and figure legend from Castelli, L. M., Cutillo, L., et al., 2021, Mol Neurodegener, PubMed 34376242.

ab26212 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab26212 please see the product overview.

Generation of whole-cell and cytoplasmic transcriptomes from healthy and C9ORF72-ALS patient-derived neurons. A Three healthy control and three C9ORF72-ALS (C9-ALS) lines of patient-derived neurons were treated with Ctrl-RNAi (C-RNAi) or SRSF1-RNAi (ΔSRSF1) prior to whole-cell (T) lysis or nuclear (N) and cytoplasmic (C) fractionation. Western blots were probed for the nuclear chromatin remodelling SSRP1 factor and the neuronal cytoplasmic marker TUJ1. B Relative expression levels of SRSF1 mRNA in whole-cell patient-derived neurons prepared in A were quantified using qRT-PCR in biological triplicates following normalization to U1 snRNA levels and to 100% for healthy neurons treated with C-RNAi (mean ± SEM; one-way ANOVA with Tukey's correction for multiple comparisons, ** : p < 0.01; N (qRT-PCR reactions) = 3). C Western blots analysis of SRSF1 protein expression in the three healthy and three C9-ALS neuron lines treated with either C-RNAi or SRSF1-RNAi. D Total, nuclear and cytoplasmic levels of intron1-spliced C9ORF72 transcripts (as measured by the exon1-exon3 junction) were quantified using qRT-PCR in biological triplicates following normalization to U1 snRNA levels and to 100% for whole-cell healthy neurons treated with C-RNAi (mean ± SEM; one-way ANOVA with Tukey's correction for multiple comparisons, NS : non-significant; N (qRT-PCR reactions) = 3). E Total, nuclear and cytoplasmic levels of unspliced C9ORF72 transcripts retaining intron1 (as measured by the exon1-intron1 junction) were quantified using qRT-PCR in biological triplicates following normalization to U1 snRNA levels and to 100% for whole-cell healthy neurons treated with C-RNAi (mean ± SEM; one-way ANOVA with Tukey's correction for multiple comparisons, NS : non-significant; *** : p < 0.001; **** : p < 0.0001; N (qRT-PCR reactions) = 3). F Whole cell transcriptome (WCT; left) and cytoplasmic transcriptome (CyT, right) sizes for quantified transcripts at gene level. G Combined WCT and CyT sizes for quantified transcripts at gene level. H Abundance of neuron (green), astrocyte (red) and oligodendrocyte (blue) markers in the human derived neurons (combined from all 24 protein-coding transcriptomes shown in Fig. 1F). I Principal component analysis (PCA) plot representing transcriptomes from patient-derived neurons, astrocytes, oligodendrocytes and fibroblasts of origin. J Principal component analysis (PCA) plot representing transcriptomes from patient-derived neurons, human post-mortem motor neurons (MN) and fibroblasts of origin

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