Specific protocols
| |
To our knowledge, customised protocols are not required for this product.
Please try the standard protocols listed below and let us know how you get on. |
| |
General protocols
Useful resources:Western blotting (WB) protocols:Immunohistochemistry (IHC) / Immunocytochemistry (ICC) protocols:Chromatin Immunoprecipitation (ChIP) protocols:Dot blot protocols:ELISA protocols:ELISPOT protocols: Flow cytometry / FACS protocols:Immunoprecipitation (IP) protocols: |
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"
![Western blot - ABCA1 antibody [HJ1] (ab66217)](/ps/datasheet/Images/66/ab66217/ab66217.bmp)
Western blot - ABCA1 antibody [HJ1] (ab66217)
All lanes : Anti-ABCA1 antibody [HJ1] (ab66217) at 1/2000 dilution
Lane 1 : Human liver lysate
Lane 2 : Human brain lysate
Lane 3 : Rat liver lysate
Lane 4 : Rat brain lysate
Lysates/proteins at 10 µg per lane.
Predicted band size : 254 kDa
![Western blot - ABCA1 antibody [HJ1] (ab66217)](/ps/datasheet/Images/66/ab66217/ab66217_1.bmp)
Western blot - ABCA1 antibody [HJ1] (ab66217)
All lanes : Anti-ABCA1 antibody [HJ1] (ab66217) at 1/2000 dilution
Lane 1 : wild type mouse bain lysate
Lane 2 : ABCA1 overexpressing mouse brain lysate
Lane 3 : ABCA1 (-/-) mouse liver lysate
Lane 4 : wild type mouse liver lysate
Lysates/proteins at 10 µg per lane.
Predicted band size : 254 kDa
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - ABCA1 antibody [HJ1] (ab66217)](/ps/datasheet/images/66/ab66217/ABCA1-Primary-antibodies-ab66217-1.jpg)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - ABCA1 antibody [HJ1] (ab66217)
ab66217 (4µg/ml) staining ABCA1 in human liver using an automated system (DAKO Autostainer Plus). Using this protocol there is moderate cell membrane staining throughout the liver parenchyma.
Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer EDTA pH 9.0 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.
![Flow Cytometry - ABCA1 antibody [HJ1] (ab66217)](/ps/datasheet/images/66/ab66217/ABCA1-Primary-antibodies-ab66217-2.jpg)
Flow Cytometry - ABCA1 antibody [HJ1] (ab66217)
Overlay histogram showing HepG2 cells stained with ab66217 (red line). The cells were fixed with methanol (5 min) and incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab66217, 2µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG2b [PLPV219] (ab91366, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a decreased signal in HepG2 cells fixed with 4% paraformaldehyde (10 min) used under the same conditions.
Please note that Abcam does not have data for use of this antibody on non-fixed cells. We welcome any customer feedback.
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ABCA1 antibody [HJ1] (ab66217)](/ps/datasheet/images/66/ab66217/ABCA1-Primary-antibodies-ab66217-12.jpg)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ABCA1 antibody [HJ1] (ab66217)
IHC image of ABCA1 staining in human liver formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab66217, 0.2µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
![ABCA1 antibody [HJ1] for Flow Cytometry in Human (66217)](/ps/datasheet/images/66/ab66217/ABCA1-Primary-antibodies-ab66217-4.jpg)
ABCA1 antibody [HJ1] for Flow Cytometry in Human (66217)
![Anti-ABCA1 antibody [HJ1] for Western blot in Human (66217)](/ps/datasheet/images/66/ab66217/ABCA1-Primary-antibodies-ab66217-8.JPG)
Anti-ABCA1 antibody [HJ1] for Western blot in Human (66217)
2
