Anti-PMP22 antibody [EPR23112-110]

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PMP22 antibody [EPR23112-110] (AB270400)

Immunohistochemical analysis of paraffin-embedded Human kidney tissue labeling PMP22 with ab270400 at 1/32000 dilution (0.018 μg/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Negative control : negative staining in human kidney (PMID : 7649472).

The section was incubated with ab270400 for 10 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins. Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PMP22 antibody [EPR23112-110] (AB270400)

Immunohistochemical analysis of paraffin-embedded Human nerve tissue labeling PMP22 with ab270400 at 1/32000 dilution (0.018 μg/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining in Schwann cells of human nerve (PMID : 7691737) is observed. The section was incubated with ab270400 for 10 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

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

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-PMP22 antibody [EPR23112-110] (AB270400)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized 293T cells transfected with a myc-tagged PMP22 expression construct. cells labelling PMP22 with ab270400 at 1/250 dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in 293T cells transfected with a myc-tagged PMP22 expression construct is observed. 2233S Myc-Tag (9B11) Mouse mAb (Alexa Fluor® 647 Conjugate) was used to counterstain tubulin at 1/100 dilution (Red). The Nuclear counterstain was DAPI (Blue).

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

• Flow Cyt

Lab

Flow Cytometry - Anti-PMP22 antibody [EPR23112-110] (AB270400)

Flow cytometric analysis of 2% paraformaldehyde fixed 0.1% Tween-20 permeabilized 293T transfected with myc-his tagged PMP22 overexpression construct cells labelling PMP22 with ab270400 at 1/600 dilution (0.1 ug)/ Right compared with a Rabbit monoclonal IgG (ab172730) / Left isotype control. A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as the secondary antibody. Cells were surface stained with isotype control or ab270400. Then fixed with 2% PFA for 10min followed by intracellularly stained with anti-myc tag conjugated to Alexa Fluor® 647.

• IP

Lab

Immunoprecipitation - Anti-PMP22 antibody [EPR23112-110] (AB270400)

PMP22 was immunoprecipitated from 0.35 mg 293T (human embryonic kidney epithelial cell) transfected with PMP22 (WT) expression vector containing a myc-His-tag®, whole cell lysate with ab270400 at 1/30 dilution (2 μg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab270400 1/1000 dilution (0.58 μg/ml). VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : 293T cells transfected with PMP22 (WT) expression vector containing a myc-His-tag®, whole cell lysate 10 μg

Lane 2 : ab270400 IP in 293T transfected with PMP22 (WT) expression vector containing a myc-His-tag®, whole cell lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab270400 in 293T transfected with PMP22 (WT) expression vector containing a myc-His-tag®, whole cell lysate. Blocking and dilution buffer and concentration : 5% NFDM/TBST.

Exposure time : 3 seconds

All lanes:

Immunoprecipitation - Anti-PMP22 antibody [EPR23112-110] (ab270400)

Predicted band size: 18 kDa

Observed band size: 25-22 kDa

false

• WB

Lab

Western blot - Anti-PMP22 antibody [EPR23112-110] (AB270400)

The expression profile observed is consistent with what has been described in the literature (PMID : 25429154; 7649472).
Negative control : Human brain (PMID : 7649472).

Blocking/diluting buffer and concentration : 5% NFDM/TBST

All lanes:

Western blot - Anti-PMP22 antibody [EPR23112-110] (ab270400) at 1/1000 dilution

Lane 1:

Human leg nerve tissue lysate at 20 µg

Lane 2:

Human brain tissue lysate at 20 µg

Secondary

All lanes:

Western blot - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/1000 dilution

Predicted band size: 18 kDa

Observed band size: 18 kDa,22 kDa

false

Exposure time: 3s

• WB

Lab

Western blot - Anti-PMP22 antibody [EPR23112-110] (AB270400)

All lanes:

Western blot - Anti-PMP22 antibody [EPR23112-110] (ab270400) at 1/1000 dilution

Lane 1:

293T (human embryonic kidney epithelial cell) transfected with an empty vector (vector control) containing a myc-His-tag®, whole cell lysate at 20 µg

Lane 2:

293T transfected with PMP22 (WT) expression vector containing a myc-His-tag®, whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/1000 dilution

Predicted band size: 18 kDa

Observed band size: 18 kDa,20 kDa

false

Exposure time: 15s