Immunofluorescence analysis of F(ab')2-Goat anti-Rabbit IgG (H+L) Secondary Antibody, Qdot 625 was performed using A431 cells stained with EGFR (EP38Y) Rabbit Monoclonal Primary Antibody (MA514485). The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, blocked with 1% BSA for 1 hour and labeled with 2ug/ml rabbit primary antibody for 3 hours at room temperature. F(ab')2-Goat anti-Rabbit IgG (H+L) Secondary Antibody, Qdot 625 (A10194) was used at a concentration of 4ug/ml in phosphate buffered saline containing 0.2 % BSA for 45 minutes at room temperature, for detection of EGFR (EP38Y) in the cytoplasm (Panel a: red). Nuclei (Panel b: blue) were stained with DAPI in SlowFade® Gold Antifade Mountant (S36938). F-actin was stained with Alexa Fluor® 488 Phalloidin (A12379, 1:300) (Panel c: green). Panel d represents the composite image. No nonspecific staining was observed with the secondary antibody alone (panel f), or with an isotype control (panel e). The images were captured at 60X magnification.
|Tested species reactivity||Rabbit|
|Published species reactivity||Not Applicable|
|Host / Isotype||Goat / IgG|
|Immunogen||Gamma Immunoglobins Heavy and Light chains|
|Storage buffer||0.05M borate, pH 8.3, with 1M betaine|
|Contains||0.05% sodium azide|
|Storage Conditions||4° C, store in dark|
|Antibody Form||F(ab')2 Fragment|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||1:50|
|Immunocytochemistry (ICC)||4 µg/ml|
|Immunofluorescence (IF)||4 µg/ml|
|Western Blot (WB)||1:50|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
|Miscellaneous PubMed (MISC)||See 1 publications below|
Qdot nanocrystals are composed of semi-conductor material to generate a fluorescent particle which is exceptionally bright and does not photobleach. Qdot nanocrystals paired with the correct optical filters are as much as 50 times brighter than traditional organic dyes.
Anti-Rabbit secondary antibodies are affinity-purified antibodies with well-characterized specificity for rabbit immunoglobulins and are useful in the detection, sorting or purification of its specified target. Secondary antibodies offer increased versatility enabling users to use many detection systems (e.g. HRP, AP, fluorescence). They can also provide greater sensitivity through signal amplification as multiple secondary antibodies can bind to a single primary antibody. Most commonly, secondary antibodies are generated by immunizing the host animal with a pooled population of immunoglobulins from the target species and can be further purified and modified (i.e. immunoaffinity chromatography, antibody fragmentation, label conjugation, etc.) to generate highly specific reagents.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
|Not Applicable||Not Cited||Quenching of biotinylated aequorin bioluminescence by dye-labeled avidin conjugates: application to homogeneous bioluminescence resonance energy transfer assays.||Adamczyk M,Moore JA,Shreder K||Organic letters (3:1797)||2001|