Anti-Clathrin heavy chain antibody

• WB

Unknown

Western blot - Anti-Clathrin heavy chain antibody (AB21679)

Blocking buffer : 2% BSA

Gel type : TA

Exposure Time : 1 minute

All lanes:

Western blot - Anti-Clathrin heavy chain antibody (ab21679) at 1 µg/mL

Lane 1:

HeLa whole cell lysate at 10 µg

Lane 2:

A431 whole cell lysate at 10 µg

Lane 3:

NIH 3T3 whole cell lysate at 10 µg

Lane 4:

PC12 whole cell lysate at 10 µg

Secondary

All lanes:

Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/50000 dilution

Predicted band size: 191 kDa

Observed band size: 180 kDa

false

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Clathrin heavy chain antibody (AB21679)

ab21679 staining Clathrin heavy chain in HeLa cells. The cells were fixed with 4% paraformaldehyde (10 min), permeabilized with 0.1% PBS-Tween for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab21679 at 1μg/ml and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control. Cells were then incubated with ab150081, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor^® 488), pre-adsorbed at 1/1000 dilution (shown in green) and ab150120, Goat polyclonal Secondary Antibody to Mouse IgG - H&L (Alexa Fluor^® 594), pre-adsorbed at 1/1000 dilution (shown in pseudocolour red). Nuclear DNA was labelled with DAPI (shown in blue).

Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.

• ICC/IF

AbReview43953****

Immunocytochemistry/ Immunofluorescence - Anti-Clathrin heavy chain antibody (AB21679)

Immunocytochemical analysis of mouse bEnd.3 labeling Clathrin heavy chain with ab21679 at 1/300 dilution using secondary anti-rabbit-alexa fluor 488 at 1/500 dilution.

This image is courtesy of an anonymous Abreview

• ICC/IF

AbReview15420****

Immunocytochemistry/ Immunofluorescence - Anti-Clathrin heavy chain antibody (AB21679)

ab21679 staining Clathrin heavy chain in mouse embryonic fibroblasts by Immunocytochemistry/ Immunofluorescence. Cells were fixed with paraformaldehyde and permeabilized with -20°C ethanol. Samples were incubated with primary antibody at 1/500 dilution (in 0.1% Saponin/1% BSA/PBS) for 1 hour. An Alexa Fluor^® 546-conjugated goat polyclonal to rabbit IgG (H+L) was used as secondary antibody at 1/500 dilution. Green color in image show positive staining with Alexa Fluor^® 546 conjugated secondary.

This image is a courtesy of Anonymous Abreview

• WB

AbReview43911****

Western blot - Anti-Clathrin heavy chain antibody (AB21679)

All lanes:

Western blot - Anti-Clathrin heavy chain antibody (ab21679) at 1/1000 dilution

All lanes:

HUVEC cell lysate at 20 µg

Secondary

All lanes:

HRP conjugated goat anti-rabbit IgG at 1/10000 dilution

Predicted band size: 191 kDa

true

Exposure time: 5s

This image is courtesy of an anonymous Abreview

• ICC/IF

CiteAb

Immunocytochemistry/ Immunofluorescence - Anti-Clathrin heavy chain antibody (AB21679)

Immunocytochemistry-immunofluorescence using Anti-Clathrin heavy chain antibody, ab21679. Publication image from Ruoslahti, E. et al., 2014, Nat Commun, 25277522. Legend direct from paper.

CendR endocytosis is mechanistically distinct from known endocytic pathways(A) The effect of knocking down selected hit genes from the genome screen on R-Ag and TFuptake using individual siRNAs format. The values represent relative probe uptakenormalized to that of negative controls (which is 1, Supplementary Data 2). The heat-map andclustering analysis were generated using Gene-E (Broad institute).(B) R-Ag uptake is not affected by Cav-ME and MP inhibitors. PPC1 cells were treated withindicated inhibitors (mβCD or Rottlerin) followed by testing for probe uptake(R-Ag, CtxB or Dex) as described in Methods. The fluorescence intensity of each probe wasnormalized to the average of the corresponding negative controls (vehicle alone) asrelative uptake (y-axis). *P<0.05 and **P<0.01(Student's t-test).(C) CendR cargo does not compete with other endocytic pathways. Unlabeled CendR peptide,RPARPAR-OH, was added to culture media of PPC1 cells at the indicated concentrations(upper right corner) 10 min prior to addition of fluorescently labeled endocytic probes(x-axis). The intensity of probe signal was normalized to the average of untreated cells(0 µM) as relative uptake (y-axis).(D) R-Ag does not colocalize with structural components of CME and Cav-ME vesicles. PPC1cells were incubated with R-Ag, TF-594 or CtxB (red) for 15 min before washing andfixation. CLTC and CAV1 proteins were detected with rabbit anti-CLTC and anti-CAV1antibodies, followed by staining with anti-rabbit secondary antibody (green). Nuclei werelabeled with Hoechst 33342 (blue). Representative images captured by confocal microscopyare shown. The colocalization events between two probes were identified with the“colocalization highlighter” macro for Image J and shown in yellow. Scalebar, 10 µm. Error bars indicate SEM (3-5 replicates).

• ICC/IF

CiteAb

Immunocytochemistry/ Immunofluorescence - Anti-Clathrin heavy chain antibody (AB21679)

Immunocytochemistry-immunofluorescence using Anti-Clathrin heavy chain antibody, ab21679. Publication image from Schmoranzer, J. et al., 2015, Nat Commun, 26399746. Legend direct from paper.

Newly exocytosed proteins co-cluster with endocytic proteins in the periactive zone.(a) 3D-Structured Illumination Microscopy (3D-SIM) analysis of Bassoon (Bass), AP180 and clathrin heavy chain (CHC) in stimulated (40 APs, 20 Hz) or non-stimulated hippocampal neurons. Scale bar, 1 µm; zoom, 500 nm. (b) Schematic for immunolabeling of the newly exocytosed SV protein pool (NEP). (c–e) SD-dSTORM imaging of newly exocytosed SV proteins (stimulation : 40 AP, 20 Hz) in hippocampal neurons expressing Syb2-c-myc. (c) 2-colour SD-dSTORM analysis of NEP and AP180, overlaid with wide-field image of Bassoon. Scale bar, 1 µm; zoom, 100 nm. (d) Ripley's L(r)-r function shows sub-synaptic clustering of NEP and of AP180, indicated by positive L(r)-r values with a maximum at radii of about 100 nm (NEP) and 200 nm (AP180). Mean±s.e.m.; n=31 synapses. Dotted lines represent upper envelopes of complete spatial randomness (CSR). (e) Radial intensity profiles of NEP and AP180 signals centred on the maxima of sub-synaptic NEP clusters show colocalization of both proteins (mean±s.e.m.; n=47 synapses). (f) k-Nearest neighbour analysis (k=10) of NEP and AP180 showing a maximum at 25 nm. The red line represents results after toroidal shift of one channel. Values above the red line indicate colocalization of NEP and AP180 (mean±s.e.m.; n=31 synapses).