LRRK2抗体Sampler组合

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - LRRK2 Antibody Sampler Panel (AB263464)

ab133475, at 1/200 dilution, staining LRRK2 in paraffin embedded Human hippocampus tissue using immunohistochemical analysis.

This image is courtesy of the Douglas Walker Lab

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - LRRK2 Antibody Sampler Panel (AB263464)

Immunofluorescent staining of SH-SY5Y cells fixed and permeablized with 4% PFA and 0.1% Triton X 100 using purified ab133518 at a dilution of 1/200. An Alexa Fluor^® 488 goat anti-rabbit was used as the secondary (ab150077, 1/400) and the sample was stained with DAPI. The negative control is shown in bottom right hand panel - for the negative control, purified ab133518 was used at a dilution of 1/200 followed by an Alexa Fluor^® 594 goat anti-mouse antibody (ab150120) at a dilution of 1/500.

• WB

Lab

Western blot - LRRK2 Antibody Sampler Panel (AB263464)

Lane 1 : Wild type A549 whole cell lysate (20 μg)
Lane 2 : Wild type MEF whole cell lysate (20 μg)
Lane 3 : LRRK2 knockout A549 whole cell lysate (20 μg)
Lane 4 : LRRK2 knockout MEF whole cell lysate (20 μg)

Lanes 1 - 4 : Merged signal (red and green). Green - Anti-LRRK2 antibody [MJFF2 (c41-2)] (ab133474) observed at 286 kDa. Red - loading control, ab18058, observed at 130 kDa.

ab133474 was shown to recognize LRRK2 in wild type A549 and MEF cells along with additional cross reative bands. Whilst signal was not seen in LRRK2 knockout cells. Wild-type and LRRK2 knockout samples were subjected to SDS-PAGE. ab133474 and ab18058 (Mouse anti Vinculin loading control) were incubated overnight at 4°C at 10000 dilution and 1/10000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

Wild-type and LRRK2 knockout MEF and A549 cells were provide as a generous gift from Professor Dario Alessi, MRC Protein Phosphorylation and Ubiquitination Unit (University of Dundee).

All lanes:

Western blot - Anti-LRRK2 antibody [MJFF2 (c41-2)] (<a href='/products/primary-antibodies/lrrk2-antibody-mjff2-c41-2-ab133474'>ab133474</a>)

Predicted band size: 286 kDa

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• WB

Supplier Data

Western blot - LRRK2 Antibody Sampler Panel (AB263464)

All lanes : Anti-LRRK2 (phospho S1292) antibody [MJFR-19-7-8] (ab203181) at 1 μg/ml

Lane 1 : GFP-LRRK2 wt transfected HEK293 lysate subjected to immunoprecipitation with GFP trap agarose

Lane 2 : GFP-LRRK2 mutant D2017A transfected HEK293 lysate subjected to immunoprecipitation with GFP trap agarose

Lane 3 : GFP-LRRK2 mutant S1292A/G2019S transfected HEK293 lysate subjected to immunoprecipitation with GFP trap agarose

Lane 4 : GFP-LRRK2 mutant G2019S transfected HEK293 lysate subjected to immunoprecipitation with GFP trap agarose

Lysates/proteins at 10 μg per lane.

Secondary

All lanes : Goat anti-rabbit (IRDye 800) at 1/10000 dilution

Predicted band size : 286 kDa

Observed band size : 313 kDa

Blocking/Dilution buffer : 5% BSA/TBST.

The image is provided by Dr. Jeremy Nicols, Parkinson's Institute, Sunnyvale, CA, USA.

G2019S mutation results in an increased LRRK2 auto-phosphorylation including S1292 (lane 4).

Observed band size : 286 + 27 (GFP) = 313 kDa.

All lanes:

Western blot - Anti-LRRK2 (phospho S1292) antibody [MJFR-19-7-8] (<a href='/products/primary-antibodies/lrrk2-phospho-s1292-antibody-mjfr-19-7-8-ab203181'>ab203181</a>) at 1 µg/mL

Lane 1:

GFP-LRRK2 wt transfected HEK293 lysate subjected to immunoprecipitation with GFP trap agarose at 10 µg

Lane 2:

GFP-LRRK2 mutant D2017A transfected HEK293 lysate subjected to immunoprecipitation with GFP trap agarose at 10 µg

Lane 3:

GFP-LRRK2 mutant S1292A/G2019S transfected HEK293 lysate subjected to immunoprecipitation with GFP trap agarose at 10 µg

Lane 4:

GFP-LRRK2 mutant G2019S transfected HEK293 lysate subjected to immunoprecipitation with GFP trap agarose at 10 µg

Secondary

All lanes:

Goat anti-rabbit (IRDye 800) at 1/10000 dilution

Predicted band size: 286 kDa

Observed band size: 313 kDa

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