Anti-Frataxin antibody [18A5DB1]

• WB

Unknown

Western blot - Anti-Frataxin antibody [18A5DB1] (AB110328)

All lanes:

Western blot - Anti-Frataxin antibody [18A5DB1] (ab110328) at 5 µg/mL

All lanes:

Recombinant Human Frataxin at 0.0005 µg

Predicted band size: 23 kDa

false

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Frataxin antibody [18A5DB1] (AB110328)

Immunohistochemical analysis of formalin fixed paraffin embedded human colon adenocarcinoma labelling frataxin with ab110328 at a concentration of 0.5µg/ml. The immunostaining was performed on a Ventana DISCOVERY ULTRA (Roche Tissue Diagnostics) instrument with a OptiView DAB IHC Detection Kit. Heat mediated antigen retrieval was performed with DISCOVERY cell conditioning solution (CC1) 100°C, pH8.5 for 32 mins. ab110328 anti-Frataxin antibody [18A5DB1] was incubated for 16 mins at 37°C. Sections were counterstained with Hematoxylin II. Image inset shows absence of staining in secondary antibody only control.

Customers are encouraged to optimise antigen retrieval conditions, antibody concentration, incubation times and temperature for best results in their own IHC assay workflow (automated and manual).

• ICC/IF

AbReview39966****

Immunocytochemistry/ Immunofluorescence - Anti-Frataxin antibody [18A5DB1] (AB110328)

Immunofluorescent analysis of paraformaldehyde-fixed, 0.2% NP-40 permeabilized SH-SY5Y (human neuroblastoma cell line from bone marrow) cells labeling Frataxin with ab110328 at 1/500 dilution, followed by Goat anti-MouseAlexaFluor^® 594 conjugated secondary antibody at 1/500 dilution (red). The nuclear counter stain is DAPI (blue).

Image courtesy of Dr George Allen

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Frataxin antibody [18A5DB1] (AB110328)

IHC image of Frataxin staining in Human colon adenocarcinoma formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab110328, 10μg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Frataxin antibody [18A5DB1] (AB110328)

Immunocytochemistry analysis using ab110328 at 0.5μg/ml staining Frataxin in normal Human fibroblasts (MRC5) (fixed and permeabilized) followed by an AlexaFluor® 594-conjugated-goat-anti-mouse IgG1 isotype specific secondary antibody (2 µg/ml). .

• Flow Cyt

Unknown

Flow Cytometry - Anti-Frataxin antibody [18A5DB1] (AB110328)

Flow cytometric analysis using ab110328 at 1µg/ml staining Frataxin in HL60 cells (blue). Isotype control antibody (red).

• WB

CiteAb

Western blot - Anti-Frataxin antibody [18A5DB1] (AB110328)

Western Blotting using Anti-Frataxin antibody [18A5DB1], ab110328. Publication image from Corey, D. R. et al., 2016, Nat Commun, 26842135. Legend direct from paper.

LNA-mediated activation of FXN expression.(a) Structure of LNA. (b) Western analysis of the effect of LNAs with phosphodiester (PO) backbone on FXN protein expression. (c) Quantitation of western analysis (n=2). (d) Western analysis of the effect of LNAs with phosphorothioate (PS) backbone on FXN protein expression. (e) Quantitation of quadruplicate western analysis. (f,g) qPCR showing effect on FXN mRNA expression of (f) PO LNAs (n=5) or (g) PS LNAs (n=3). PO-control-LNA5 is a negative control LNA with PO backbone that is not complementary to FXN RNA. PO-control-LNA6 has five mismatches relative to the repeat region within FXN RNA target. PS-control-LNA7 is a negative control LNA similar to PO-control-LNA5 but with PS backbone. FRDA patient fibroblast cells (GM03816) were treated with 12.5 nM duplex LNAs. Cells were collected at day 3 for RNA extraction and day 4 for protein extraction. All data are presented as mean±STDEV. NT, no treatment; **P<0.01, by Student t-test.

false

• WB

CiteAb

Western blot - Anti-Frataxin antibody [18A5DB1] (AB110328)

Western Blotting using Anti-Frataxin antibody [18A5DB1], ab110328. Publication image from Corey, D. R. et al., 2016, Nat Commun, 26842135. Legend direct from paper.

RNA-mediated activation of FXN expression.(a) Schematic of repeat expansion within intronic FXN mRNA and binding of AGO : RNA complexes. The longer mutant repeat is predicted to bind more AGO : RNA complexes than the shorter wild-type repeat. (b) Schematic of R-loop formation at FXN locus and potential to influence histone modification and gene expression (Adapted from Groh et al.19). (c) Complementarity of guide strand RNAs (25 nM) to FXN RNA. (d,e) Effect of anti-GAA duplex RNAs on d, FXN mRNA (n=3) and e, protein expression. (f) A dose-response profile of upregulation of FXN protein expression by siGAA. FRDA patient-derived fibroblast cells (GM03816) were used. siExon2 is a duplex siRNA that targets FXN exon and is expected to decrease FXN expression. CM is a negative control RNA that is not complementary to FXN RNA. RNA scramble is a duplex RNA in which the sequences of siGAA and siAAG are mixed to preserve nucleotide composition by alter their order. si5mmAAG is similar to siAAG but has five mismatches relative to the repeat region within FXN mRNA target. Cells were collected at day 3 for RNA extraction and day 4 for protein extraction. Error bars : ±STDEV. **P<0.01, by Student t-test.

false

• WB

CiteAb

Western blot - Anti-Frataxin antibody [18A5DB1] (AB110328)

Western Blotting using Anti-Frataxin antibody [18A5DB1], ab110328. Publication image from Corey, D. R. et al., 2016, Nat Commun, 26842135. Legend direct from paper.

Comparison of RNA-mediated activation of FXN expression in patient cells to FXN expression in normal cells and FXN activation by a histone deacetylase inhibitor.(a) Activation of FXN protein expression in FRDA cells (GM03816) and wild-type fibroblast cells (GM02153). (b,c) Effect of HDAC inhibitor BML210 (5 µM) treatment on expression of bFXN mRNA (n=3) and (c) protein (inset, western analysis, n=4) expression in FRDA patient fibroblast cells (GM03816). All data are presented as mean±STDEV.

false

• WB

CiteAb

Western blot - Anti-Frataxin antibody [18A5DB1] (AB110328)

Western Blotting using Anti-Frataxin antibody [18A5DB1], ab110328. Publication image from Corey, D. R. et al., 2016, Nat Commun, 26842135. Legend direct from paper.

Comparison of RNA-mediated activation of FXN expression in patient cells to FXN expression in normal cells and FXN activation by a histone deacetylase inhibitor.(a) Activation of FXN protein expression in FRDA cells (GM03816) and wild-type fibroblast cells (GM02153). (b,c) Effect of HDAC inhibitor BML210 (5 µM) treatment on expression of bFXN mRNA (n=3) and (c) protein (inset, western analysis, n=4) expression in FRDA patient fibroblast cells (GM03816). All data are presented as mean±STDEV.

false