Anti-LY6C antibody [RM1151] - BSA and Azide free

• mIHC

Supplier Data

Multiplex immunohistochemistry - Anti-LY6C antibody [RM1151] - BSA and Azide free (AB317273)

This data was developed using ab317272, the same antibody clone in a different buffer formulation.

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded mouse spleen tissue staining LY6C with ab317272 at a 1 : 100 (0.978 ug/ml) dilution, ab133357 anti-CD11b used at 1 : 20000 (0.05 ug/ml) dilution and ab245235 anti-CD19 used at a 1 : 1000 (0.999 ug/ml) dilution.

Panel A : merged staining of anti-Ly6c (green; Opal™690), anti-CD11b (magenta; Opal™570) and anti-CD19 (yellow; Opal™520) on mouse spleen.
Panel B : anti-Ly6c stained on monocytes/macrophages.
Panel C : anti-CD11b stained on monocytes/macrophages.
Panel D : anti-CD19 stained on B cells.
Co-staining of Ly6c and CD11b can be observed.
The section was incubated in three rounds of staining : in the order of ab317272, ab133357, and ab245235 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.
Nuclear DNA was labeled with DAPI (shown in blue).

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

Nuclear counter stain with DAPI.

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LY6C antibody [RM1151] - BSA and Azide free (AB317273)

This data was developed using ab317272, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Mouse colon tissue labeling LY6C with ab317272 at 1/500 (0.978 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on endothelial cells and immune cells in mouse colon. The section was incubated with ab317272 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LY6C antibody [RM1151] - BSA and Azide free (AB317273)

This data was developed using ab317272, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Mouse cerebrum tissue labeling LY6C with ab317272 at 1/500 (0.978 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on endothelial cells in mouse cerebrum. The section was incubated with ab317272 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LY6C antibody [RM1151] - BSA and Azide free (AB317273)

This data was developed using ab317272, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Mouse spleen tissue labeling LY6C with ab317272 at 1/500 (0.978 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on endothelial cells and immune cells in mouse spleen. The section was incubated with ab317272 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-LY6C antibody [RM1151] - BSA and Azide free (AB317273)

This data was developed using ab317272, the same antibody clone in a different buffer formulation.

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized Mouse splenocyte cells labelling LY6C with ab317272 at 1/500 (0.978 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2 ug/ml) dilution (Green).

Confocal image showing cytoplasmic staining in subsets of mouse splenocytes (shown in green), which is partially co-located with CD11b. The counterstain was observed in magenta. Nuclear DNA was labeled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab8878 Anti-CD11b Rat monoclonal antibody was used to counterstain at 1/200 (2.5ug/ml) dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 (2 ug/ml) dilution.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-LY6C antibody [RM1151] - BSA and Azide free (AB317273)

This data was developed using ab317272, the same antibody clone in a different buffer formulation.

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized Mouse bone marrow cells labelling LY6C with ab317272 at 1/500 (0.978 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2 ug/ml) dilution (Green).

Confocal image showing cytoplasmic staining in subsets of mouse bone marrow (shown in green), which is partially co-located with CD11b. The counterstain was observed in magenta. Nuclear DNA was labeled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab8878 Anti-CD11b Rat monoclonal antibody was used to counterstain at 1/200 (2.5ug/ml) dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 (2 ug/ml) dilution.

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-LY6C antibody [RM1151] - BSA and Azide free (AB317273)

This data was developed using ab317272, the same antibody clone in a different buffer formulation.

Flow cytometric analysis of Mouse bone marrow cells labelling LY6C with ab317272 at 1/500 dilution (0.1 ug)/Right compared with a Rabbit monoclonal IgG (ab172730) / Left isotype control.

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

Mouse bone marrow are co-stained with CD11b conjugated BV510 (512/25BP).
Gated on viable cell.

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-LY6C antibody [RM1151] - BSA and Azide free (AB317273)

This data was developed using ab317272, the same antibody clone in a different buffer formulation.

Flow cytometric analysis of Mouse spleen cell cells labelling LY6C with ab317272 at 1/500 dilution (0.1 ug)/Right compared with a Rabbit monoclonal IgG (ab172730) / Left isotype control.

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

Mouse spleen cell are co-stained with CD8a conjugated Alexa Fluor®647.
Gated on viable cell.

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-LY6C antibody [RM1151] - BSA and Azide free (AB317273)

This data was developed using ab317272, the same antibody clone in a different buffer formulation.

Flow cytometric analysis of Mouse bone marrow cells labelling LY6C with ab317272 at 1/500 dilution (0.1 ug)/Right compared with a Rabbit monoclonal IgG (ab172730) / Left isotype control.

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

Mouse bone marrow are co-stained with B220 conjugated Alexa Fluor®647.
Gated on viable cell.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LY6C antibody [RM1151] - BSA and Azide free (AB317273)

This data was developed using ab317272, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Mouse liver tissue labeling LY6C with ab317272 at 1/500 (0.978 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Low expression tissue : no staining on mouse liver. The section was incubated with ab317272 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IP

Supplier Data

Immunoprecipitation - Anti-LY6C antibody [RM1151] - BSA and Azide free (AB317273)

This data was developed using ab317272, the same antibody clone in a different buffer formulation.

LY6C was immunoprecipitated from 0.35 mg Mouse spleen tissue lysate with ab317272 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab317272 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : Mouse spleen tissue lysate
Lane 2 : ab317272 IP in Mouse spleen tissue lysate
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab317272 in mouse spleen tissue lysate.

All lanes:

Immunoprecipitation - Anti-LY6C antibody [RM1151] (<a href='/en-us/products/primary-antibodies/ly6c-antibody-rm1151-ab317272'>ab317272</a>) at 1/30 dilution

All lanes:

Mouse spleen tissue lysate with 5% NFDM/TBST

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

false

Exposure time: 128s

• WB

Supplier Data

Western blot - Anti-LY6C antibody [RM1151] - BSA and Azide free (AB317273)

This data was developed using ab317272, the same antibody clone in a different buffer formulation.

Negative control : Neuro-2a.

Low expression : testis, liver.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-LY6C antibody [RM1151] (<a href='/en-us/products/primary-antibodies/ly6c-antibody-rm1151-ab317272'>ab317272</a>) at 1/1000 dilution

Lane 1:

Mouse lung tissue lysate at 20 µg with 5% NFDM/TBST

Lane 2:

Mouse spleen tissue lysate at 20 µg with 5% NFDM/TBST

Lane 3:

Mouse testis tissue lysate at 20 µg with 5% NFDM/TBST

Lane 4:

Mouse liver tissue lysate at 20 µg with 5% NFDM/TBST

Lane 5:

Mouse kidney tissue lysate at 20 µg with 5% NFDM/TBST

Lane 6:

NIH/3T3 (mouse embryonic fibroblast) whole cell lysate at 20 µg with 5% NFDM/TBST

Lane 7:

C2C12 (mouse myoblast) whole cell lysate at 20 µg with 5% NFDM/TBST

Lane 8:

Neuro-2a (mouse neuroblastoma neuroblast) whole cell lysate at 20 µg with 5% NFDM/TBST

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

Observed band size: 14 kDa,36 kDa

false

Exposure time: 180s