Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Mouse cerebral cortex tissue sections labeling Tyrosine Hydroxylase with Purified ab137869 at 1:500 dilution (1.1 μg/ml). Heat mediated antigen retrieval was performed using Perform heat mediated antigen retrieval using citrate Buffer, PH6. Tissue was counterstained with Hematoxylin. ab97051 Goat Anti-Rabbit IgG H&L (HRP) secondary antibody was used at 1:500 dilution. PBS instead of the primary antibody was used as the negative control.

Immunocytochemistry/ Immunofluorescence analysis of C6 (Rat glial tumor cell line) cells labeling Tyrosine Hydroxylase with Purified ab137869 at 1:100 dilution (5.6μg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) 1:200 (2.5 μg/ml). ab150077 Goat anti rabbit IgG(Alexa Fluor^® 488) was used as the secondary antibody at 1:1000 dilution. DAPI nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

Flow Cytometry analysis of SH-SY5Y (Human neuroblastoma cell line from bone marrow) cells labeling Tyrosine Hydroxylase with purified ab137869 at 1:50 dilution (10 ug/ml) (red). Cells were fixed with 4% Paraformaldehyde. A Goat anti rabbit IgG (Alexa Fluor^® 488) secondary antibody was used at 1:2000 dilution. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

All lanes : Anti-Tyrosine Hydroxylase antibody [EP1532Y] (ab137869) at 0.03 µg/ml (purified)

Lane 1 : Mouse brain lysate
Lane 2 : Rat brain lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Predicted band size: 58 kDa
Observed band size: 58 kDa

Blocking and diluting buffer: 5% NFDM/TBST

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Rat cerebral cortex tissue sections labeling Tyrosine Hydroxylase with Purified ab137869 at 1:500 dilution (1.1 μg/ml). Heat mediated antigen retrieval was performed using Perform heat mediated antigen retrieval using citrate Buffer, PH6. Tissue was counterstained with Hematoxylin. ab97051 Goat Anti-Rabbit IgG H&L (HRP) secondary antibody was used at 1:500 dilution. PBS instead of the primary antibody was used as the negative control.

ab137869 staining Tyrosine Hydroxylase in mouse brain tissue sections by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue was fixed with formaldehyde and a heat mediated antigen retrieval step was performed using citrate buffer. Samples were then blocked with 1% B.S.A. for 10 minutes at 21ºC followed by incubation with the primary antibody for 2 hours at 1/800. A biotin-conjugated goat anti-rabbit polyclonal was used as secondary antibody at a 1/250 dilution.

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ab137869 staining Tyrosine Hydroxylase in rat brain tissue sections by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue was fixed with formaldehyde and a heat mediated antigen retrieval step was performed using citrate buffer. Samples were then blocked with 1% B.S.A. for 10 minutes at 21ºC followed by incubation with the primary antibody for 2 hours at 1/1000. A biotin-conjugated goat anti-rabbit polyclonal was used as secondary antibody at a 1/250 dilution.

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Anti-Tyrosine Hydroxylase antibody [EP1532Y] (ab137869) at 0.1 µg/ml (purified) + Human adrenal gland lysate at 20 µg

Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Predicted band size: 58 kDa
Observed band size: 58 kDa

Blocking and diluting buffer : 5% NFDM/TBST

Overlay histogram showing SHSY-5Y cells stained with ab137869 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab137869, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1μg/1x10⁶ cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. This antibody gave a positive signal in SHSY-5Y cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

Anti-Tyrosine Hydroxylase antibody [EP1532Y] (ab137869) at 1/100000 dilution (unpurified) + PC12 cell lysate at 10 µg

Secondary
Goat anti-rabbit HRP at 1/2000 dilution

Predicted band size: 58 kDa

All lanes : Anti-Tyrosine Hydroxylase antibody [EP1532Y] (ab137869) at 1/5000 dilution (unpurified)

Lane 1 : SH-SY5Y cell lysate - transduced with AAV vector expressing human TH
Lane 2 : SH-SY5Y cell lysate - non-infected

Lysates/proteins at 20000 cells per lane.

Secondary
All lanes : HRP-conjugated goat anti-rabbit IgG at 1/10000 dilution

Developed using the ECL technique.

Performed under non-reducing conditions.

Predicted band size: 58 kDa
Observed band size: 58 kDa


Exposure time: 1 second

Blocked with 5% milk for 1 hour at 25°C.

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