Anti-Niemann Pick C1 antibody [EPR5209] - BSA and Azide free

• ICC/IF

PubMed

Immunocytochemistry/ Immunofluorescence - Anti-Niemann Pick C1 antibody [EPR5209] - BSA and Azide free (AB224268)

Cathepsin B/L inhibition causes NPC disease-like cholesterol accumulation in SH-SY5Y cells.

Confocal microscopy of SH-SY5Y control and PADK treated cells. Cholesterol (filipin staining, white) and NPC1 (ab134113; green).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

Image from Cermak S et al., PLoS One. 2016;11(11):e0167428. Fig 3.; doi: 10.1371/journal.pone.0167428. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Niemann Pick C1 antibody [EPR5209] - BSA and Azide free (AB224268)

Immunohistochemical staining of paraffin embedded human liver with purified ab134113 at a working dilution of 1 in 50. The secondary antibody used is a HRP polymer for rabbit IgG. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Niemann Pick C1 antibody [EPR5209] - BSA and Azide free (AB224268)

Immunohistochemical analysis of paraffin embedded human kidney tissue labelling Niemann Pick C1 with unpurified ab134113 at 1/50.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Niemann Pick C1 antibody [EPR5209] - BSA and Azide free (AB224268)

This ICC data was generated using the same anti-Niemann Pick C1 antibody clone [EPR5209] in a different buffer formulation (cat# ab134133).

Immunofluorescence staining of neuro-2a cells with purified ab134113 at a working dilution of 1 in 70, counter-stained with DAPI. The secondary antibody was Alexa Fluor^® 488 goat anti rabbit (ab150077), used at a dilution of 1 in 500. The cells were fixed in 4% PFA and permeabilized using 0.1% Triton X 100. The negative control is shown in bottom right hand panel - for the negative control, purified ab134113 was used at a dilution of 1/200 followed by an Alexa Fluor^® 594 goat anti-mouse antibody (ab150120) at a dilution of 1/500.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Niemann Pick C1 antibody [EPR5209] - BSA and Azide free (AB224268)

Immunohistochemical staining of paraffin embedded mouse liver with purified ab134113 at a working dilution of 1 in 50. The secondary antibody used is a HRP polymer for rabbit IgG. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-Niemann Pick C1 antibody [EPR5209] - BSA and Azide free (AB224268)

ab134113 staining Niemann Pick C1 in Neuro-2a (mouse neuroblastoma cell line) by intracellular flow cytometry. Cells were fixed with 4% paraformaldehyde, permabilised with 90% methanol and the sample was incubated with the primary antibody at a dilution of 1/200. A goat anti rabbit IgG (Alexa Fluor^® 488) at a dilution of 1/2000 was used as the secondary antibody.

Isoytype control : Rabbit monoclonal IgG (Black)

Unlabeled control : Cell without incubation with primary antibody and secondary antibody (Blue)

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Niemann Pick C1 antibody [EPR5209] - BSA and Azide free (AB224268)

Immunohistochemical staining of paraffin embedded rat cerebellum with purified ab134113 at a working dilution of 1 in 50. The secondary antibody used is a HRP polymer for rabbit IgG. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

• WB

Lab

Western blot - Anti-Niemann Pick C1 antibody [EPR5209] - BSA and Azide free (AB224268)

Lanes 1 - 4 : Merged signal (red and green). Green - ab134113 observed at 180 kDa. Red - loading control, ab9484, observed at 37 kDa.

ab134113 was shown to specifically react with Niemann Pick C1 in wild-type HAP1 cells as signal was lost in NPC1 (Niemann Pick C1) knockout cells. Wild-type and NPC1 (Niemann Pick C1) knockout samples were subjected to SDS-PAGE. ab134113 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab134113).

All lanes:

Western blot - Anti-Niemann Pick C1 antibody [EPR5209] (<a href='/en-us/products/primary-antibodies/niemann-pick-c1-antibody-epr5209-ab134113'>ab134113</a>) at 1/1000 dilution

Lane 1:

Wild-type HAP1 whole cell lysate at 20 µg

Lane 2:

NPC1 (Niemann Pick C1) knockout HAP1 whole cell lysate at 20 µg

Lane 3:

HEK293 whole cell lysate at 20 µg

Lane 4:

HepG2 whole cell lysate at 20 µg

Predicted band size: 142 kDa

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• OI-RD Scanning

Unknown

OI-RD Scanning - Anti-Niemann Pick C1 antibody [EPR5209] - BSA and Azide free (AB224268)

We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen), of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody. Based on the comparison with published literature values for mouse monoclonal antibodies, Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.