This site is for customers in Asia. Customers in China & other regions, please go to Global page.
HOME > Support > Principle and method of the experiment > Labeled antibodies

Labeled antibodies

Labels for detecting antibodies

Antibodies are labeled with various substances for detection.
When using a commercial kit to label antibodies, care must be taken not to lose the activity of the antibody.

Common types of labels

Fluoresecent labeling
Enzyme labeling
Biotin
Magnetic beads, agarose beads, magnetic agarose beads
Colloidal gold

Fluoresecent labeling

Fluorescent labeled antibodies are used for flow cytometry and immunofluorescence staining. Multiple antibodies with different fluorescent labels can be used for double or triple staining.

Common fluorophores
Type of fluorophore
Fluorescent dye (low molecular weight fluorescent dye) FITC (fluorescein isothiocyanate), Alexa Fluor® dye Cy dyes, etc.
Fluorescent protein PE (phycoerythrin), APC (allophycocyanin), etc.



Application example of fluorescent-labeled antibody: Immunofluorescence staining

Immunofluorescence staining with an antibody to an organelle marker
HeLa cells were stained with an antibody to the Golgi protein GM130.
Application example of fluorescent-labeled antibody: Immunofluorescence staining
Antibody:Anti-GM130 mAb-Alexa Fluor® 488(Code No. M179-A48

Application example of fluorescent-labeled antibody: Flow cytometry

T cells are separated by type using fluorescent-labeled antibodies to cell surface markers.
T cells are separated into helper T cells, cytotoxic T cells, etc., by flow cytometry using antibodies labeled with different fluorophores (CD4-FITC, CD8-PE).
Overview of FCM
Application example of fluorescent-labeled antibody: Flow cytometry
PE-labeled anti-CD8 antibody (Clone: KT15, Code No. D271-5)
FITC-labeled anti-CD4 antibody (Clone: GK1.5, Code No. D341-4)



Enzyme labeling

Enzyme-labeled antibodies are used for ELISA, Western blotting, and immunostaining.
Labeled antibodies are detected by reaction with a substrate that emits light or changes color.

Commonly used enzymes
Enzyme name Chromogenic substrate Chemiluminescence substrate
HRP(Horseradish peroxidase) DAB and TMB Luminol-based (ECL)
AP(Alkaline phosphatase) BCIP/NBT and pPNPP Dioxetane-based (CDP-star®)



Application example of enzyme-labeled antibodies: Immunohistochemical staining

Application example of enzyme-labeled antibodies: Immunohistochemical staining
The result of immunohistochemical staining indicates that autophagy was induced in the liver tissue.
Primary antibody: Anti-LC3 polyclonal antibody (Code No. PM036)
Secondary antibody: Histostar™ (Rb) for Human tissue (Peroxidase-labeled, Code No. 8466)
Substrate: DAB Substrate Solution (Histostar™) (Code No. 8469)

Application example of enzyme-labeled antibodies: Western blotting

Application example of enzyme-labeled antibodies: Western blotting
Primary antibody: Mouse anti-GFP antibody (Clone: 1E4, Code No. M048-3)
Secondary antibody: Anti-mouse IgG antibody (HRP-labeled, Code No. 330)



Biotin

Biotin-labeled antibodies are used in flow cytometry, immunofluorescence staining, and immunohistochemical staining. Biotin-labeled antibodies are detected by reaction with dye-, enzyme-, or fluorophore-labeled avidin.
Biotin labeling
Antibody: Biotin-labeled anti-GP2 (Glycoprotein 2) antibody (Code No. D278-6)


Agarose, magnetic beads, magnetic agarose beads

Antibody-conjugated resins, such as magnetic beads and agarose, are used for immunoprecipitation.
Beads labeled antibody
>>The principle and method of immunoprecipitation (IP)



Colloidal gold

Gold-labeled antibodies are used in immunoelectron microscopy. This method is highly sensitive, and the signal does not fade as in fluorescent-labeling and enzyme-labeling.

Immunoelectron micrograph of autophagosomes detected using the marker LC3
Colloidal gold: Immunoelectron micrograph
Primary antibody: Anti-LC3 monoclonal antibody (Clone: 4E12, Code No. M152-3)











Related links

Fractionation and purification of proteins