Anti-ICAM1 antibody [EPR22161-284]

• WB

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Western blot - Anti-ICAM1 antibody [EPR22161-284] (AB222736)

Blocking/Diluting buffer and concentration : 5% NFDM/TBST.

Exposure time : 48 seconds.

The 110-KDa apparent molecular mass is due to glycosylation (PMID : 1680698).

All lanes:

Western blot - Anti-ICAM1 antibody [EPR22161-284] (ab222736) at 1/1000 dilution

Lane 1:

Mouse kidney tissue lysate at 20 µg

Lane 2:

Mouse spleen tissue lysate at 20 µg

Lane 3:

Mouse liver tissue lysate at 20 µg

Lane 4:

A20 (mouse reticulum sarcoma b lymphocyte) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 57 kDa

Observed band size: 110 kDa

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• Flow Cyt

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Flow Cytometry - Anti-ICAM1 antibody [EPR22161-284] (AB222736)

Flow cytometric analysis of Mouse primary splenocytes labeling ICAM1 with ab222736 at 1/500 dilution (Red) compared with a Rabbit monoclonal IgG (ab172730) isotype control (Black) and an unlabeled control (Cell without incubation with primary antibody and secondary antibody, Blue). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/200 dilution was used as the secondary antibody. Gated on viable cells.

ICAM1 negative population (left half of red peak) was also observed in mouse splenocytes which is consistent with IHC staining pattern.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ICAM1 antibody [EPR22161-284] (AB222736)

Immunohistochemical analysis of paraffin-embedded mouse spleen tissue labeling ICAM1 using ab222736 at 1/1000 dilution, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Cytoplasmic staining on the marginal zone of mouse spleen (PMID : 12130787; PMID : 17947659) is observed. Counterstained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) ready to use.

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.

Immunostaining was performed on a Leica Biosystems BOND^® RX instrument. The section was incubated with ab222736 for 30 mins at room temperature.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-ICAM1 antibody [EPR22161-284] (AB222736)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized A20 (mouse reticulum sarcoma cell line) cells labeling ICAM with ab222736 at 1/50 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing membranous staining in A20 cells with distinct staining of the target protein being transported from Golgi to cell surfaces (PMID : 17486117) is observed. Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) was used as a countertsain (Red) at 1/200 dilution. The nuclear counterstain is DAPI (Blue).

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Alexa Fluor^®488 Goat anti-Rabbit secondary (ab150077) at 1/1000 dilution.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ICAM1 antibody [EPR22161-284] (AB222736)

Immunohistochemical analysis of paraffin-embedded mouse lung tissue labeling ICAM1 using ab222736 at 1/1000 dilution, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Membranous and cytoplasmic staining on mouse lung (PMID : 14568945; PMID : 9802985) is observed. Counterstained with DAPI.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) ready to use.

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.

Immunostaining was performed on a Leica Biosystems BOND^® RX instrument. The section was incubated with ab222736 for 30 mins at room temperature.

• IP

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Immunoprecipitation - Anti-ICAM1 antibody [EPR22161-284] (AB222736)

ICAM1 was immunoprecipitated from 0.35 mg Mouse spleen lysate with ab222736 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab222736 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1/5000 dilution.

Lane 1 : Mouse spleen tissue lysate 10 μg (input).
Lane 2 : ab222736 IP in mouse spleen tissue lysate.
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab222736 in Mouse spleen lysate.

Blocking and dilution buffer and concentration : 5% NFDM/TBST.
Exposure time : 3 minutes.

110-kDa molecular mass is due to glycosylation (PMID : 1680698).

All lanes:

Immunoprecipitation - Anti-ICAM1 antibody [EPR22161-284] (ab222736)

Predicted band size: 57 kDa

Observed band size: 110 kDa

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