重组Anti-Rhodopsin抗体[EPR21876]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Rhodopsin antibody [EPR21876] (AB221664)

Immunohistochemical analysis of paraffin-embedded human retina tissue labeling Rhodopsin with ab221664 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining in outer segment of human retina is observed (PMID : 23223288). Counter stained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• mIHC

Lab

Multiplex immunohistochemistry - Anti-Rhodopsin antibody [EPR21876] (AB221664)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded Human retina tissue labeling RPE65, Rhodopsin and Synapsin I with ab231782 at 1/8000 dilution, ab232934 at 1/8000 dilution and ab274430 at 1/1500 dilution followed by a ready to use Opal Polymer HRP Ms + Rb secondary antibody. Nuclear counter stain used was DAPI. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins Panel A : merged staining of anti-Synapsin I (magenta; Opal™690), anti-RPE65 (green; Opal™520) and anti-Rhodopsin (red; Opal™570) on human retina. Panel B : anti-RPE65 stained on pigmented layer. Panel C : anti-Rhodopsin stained on rod photoreceptor cells. Panel D : anti-Synapsin I stained on inner plexiform layer. The section was incubated in three rounds of staining : in the order of ab274430, ab231782, and ab232934 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. 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. The section was incubated in three rounds of staining : in the order of ab312840, ab16669, and ab236434 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. 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.

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-Rhodopsin antibody [EPR21876] (AB221664)

Immunohistochemical analysis of 4% paraformaldehyde-fixed, 0.2% Triton X-100 permeabilized frozen rat retina tissue labeling Rhodopsin with ab221664 at 1/500 dilution,followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Positive staining in the outer segment of rat retina is observed (PMID : 21938483).

The nuclear counter stain is DAPI (blue).

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

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-Rhodopsin antibody [EPR21876] (AB221664)

Immunohistochemical analysis of 4% paraformaldehyde-fixed, 0.2% Triton X-100 permeabilized frozen mouse retina tissue labeling Rhodopsin with ab221664 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Positive staining in the outer segment of mouse retina is observed (PMID : 21938483).

The nuclear counter stain is DAPI (blue).

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

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Rhodopsin antibody [EPR21876] (AB221664)

Immunohistochemical analysis of paraffin-embedded mouse retina tissue labeling Rhodopsin with ab221664 at 1/32000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining in outer segment of mouse retina is observed (PMID : 23223288). Counter stained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• mIHC

Lab

Multiplex immunohistochemistry - Anti-Rhodopsin antibody [EPR21876] (AB221664)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded mouse retina tissue staining RPE65 with ab231782 at a 1/8000 dilution, ab109233 anti-PAX6 used at 1/1000 dilution and ab221664 anti-RHO used at a 1/30000 dilution.

Panel A : merged staining of anti-RPE65 (green; Opal™520), anti-PAX6 (magenta; Opal™690) and anti-RHO (gray; Opal™570) on mouse retina.

Panel B : anti-RPE65 staining pigmented layer in mouse retina.

Panel C : ant-PAX6 staining inner nuclear layer and ganglion cell layer in mouse retina.

Panel D : ant-RHO staining rod-shaped photoreceptor cells in mouse retina.

Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab231782, ab109233 and ab221664 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

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.

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

• mIHC

Lab

Multiplex immunohistochemistry - Anti-Rhodopsin antibody [EPR21876] (AB221664)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded mouse retina tissue staining RPE65 with ab231782 at a 1/8000 dilution, ab140603 anti-CPX used at 1/2000 dilution and ab221664 anti-RHO used at a 1/30000 dilution.

Panel A : merged staining of anti-RPE65 (green; Opal™520), anti-CPX (magenta; Opal™690) and anti-RHO (gray; Opal™570) on mouse retina.

Panel B : anti-RPE65 staining pigmented layer in mouse retina.

Panel C : ant-CPX staining outer nuclear layer and inner nuclear layer in mouse retina.

Panel D : ant-RHO staining rod-shaped photoreceptor cells in mouse retina.

Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab231782, ab140603 and ab221664 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

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.

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-Rhodopsin antibody [EPR21876] (AB221664)

Immunohistochemical analysis of paraffin-embedded rat retina tissue labeling Rhodopsin with ab221664 at 1/32000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining in outer segment of rat retina is observed (PMID : 23223288). Counter stained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IP

Supplier Data

Immunoprecipitation - Anti-Rhodopsin antibody [EPR21876] (AB221664)

Rhodopsin was immunoprecipitated from 0.35 mg rat eye tissue lysate with ab221664 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab221664 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1/5000 dilution.

Lane 1 : Rat eye tissue lysate 10 μg (input).

Lane 2 : Rat eye tissue lysate (boiled) 10 μg (input).

Lane 3 : ab221664 IP in Rat eye tissue lysate.

Lane 4 : Rabbit monoclonal IgG (ab172730) instead of ab221664 in rat eye tissue lysate.

Exposure time : 15 seconds

The multiple bands (>60 kDa) correspond to dimers and multimers of rhodopsin, consistent with the literature (PMID : 25270370; PMID : 22219383).

We do not recommend boiling samples in loading buffer as this may cause protein aggregation (lane 2).

All lanes:

Immunoprecipitation - Anti-Rhodopsin antibody [EPR21876] (ab221664)

Predicted band size: 38 kDa

false

• WB

Supplier Data

Western blot - Anti-Rhodopsin antibody [EPR21876] (AB221664)

Exposure Time : Lane 1 : 3 minutes; Lane 2 : 3 seconds; Lane 3 : 58 seconds; Lane 4 : 26 seconds.

Blocking/Dilution buffer : 5% NFDM/TBST.

The multiple bands (>60 kDa) correspond to dimers and multimers of rhodopsin, consistent with the literature (PMID : 25270370; PMID : 22219383).

Lanes 1 - 3:

Western blot - Anti-Rhodopsin antibody [EPR21876] (ab221664) at 1/1000 dilution

Lane 4:

Western blot - Anti-Rhodopsin antibody [EPR21876] (ab221664) at 1/5000 dilution

Lane 1:

Human eye tissue at 20 µg

Lane 2:

Mouse eye tissue at 20 µg

Lane 3:

Rat retina tissue lysate at 20 µg

Lane 4:

Rat eye tissue lysate at 10 µg

Secondary

All lanes:

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

Predicted band size: 38 kDa

Observed band size: 35-150 kDa

false