All lanes : Anti-Doublecortin antibody - Neuronal Marker (ab23542) at 1 µg/mlLane 1 : Mouse brain lysateLane 2 : Mouse day 0 brain lysateLysates/proteins at 20 µg per lane.SecondaryGoat polyclonal to rabbit IgG (Alexa fluor 680) at 1/10000 dilution
All lanes : Anti-Doublecortin antibody - Neuronal Marker (ab23542) at 1 µg/mlLane 1 : Spinal Cord (Mouse) Tissue Lysate Lane 2 : Brain (Rat) Tissue Lysate - normal tissue Lysates/proteins at 10 µg per lane.SecondaryIRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilutionPerformed under reducing conditions.
ICC/IF image of ab23542 stained PC12 cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab23542, 10µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899 Dylight 488 goat anti-rabbit IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
![Doublecortin was immunoprecipitated using 0.5mg Rat brain tissue, 5µg of Rabbit polyclonal to Doublecortin and 50µl of protein G magnetic beads (+). No antibody was added to the control (-). The antibody was incubated under agitation with Protein G beads for 10min, Rat brain tissue lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab23542.Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).Band: 50kDa; Doublecortin](http://www.bioprodhub.com/system/product_images/ab_products/2/sub_2/9832_Doublecortin-Primary-antibodies-ab23542-102.jpg)
Doublecortin was immunoprecipitated using 0.5mg Rat brain tissue, 5µg of Rabbit polyclonal to Doublecortin and 50µl of protein G magnetic beads (+). No antibody was added to the control (-). The antibody was incubated under agitation with Protein G beads for 10min, Rat brain tissue lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab23542.Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).Band: 50kDa; Doublecortin