How to Select an Antibody&Protein Coupling Kit with Super Cost Performance

1, The common definition of coupling technology

Enzymes, biotin, fluorescent dye, agarose, magnetic beads and other substances which are easy to detect or separate are covalently labeled or conjugated to antibodies, proteins or other small molecular substances by a certain technology.

2, Background of Coupling Technology

Coupling of antibodies, proteins or other small molecules is a necessary step in many types of biological experiments.

For example, in order to detect specific proteins in immunoassays (such as WB/IHC/IF and other immunological experiments), we couple primary antibody or secondary antibody with enzyme, fluorescent dye, etc.

In cell staining (biological imaging) applications, reactive dyes (fluorescent dye) are directly combined with extracellular or intracellular proteins (polypeptides) to produce visible and measurable staining.

Coupling agarose or magnetic beads to proteins or antibodies provides a targeted purification method. For example, the agarose (magnetic beads) coupled antibody used by Abbkine in IP experiments directly saves the use of Protein A/G.

3, Major Disclosure of Coupling Kits of Various Brands (Taking HRP Coupling Kits and Green Fluorescence Coupling Kits as Examples)

1. HRP Coupling Kits

Brand	Sample types	Contents of the kit	Duration of experiment	Size/Price
Abbkine	Samples contain free amino group	1. Activated HRP conjugates solution 2. HRP labeling solution 3. HRP quencher	3.5-4h	3*20ug/$100 100ug/$120 3*100ug/$140 1mg/$220
Abcam	Samples contain free amino group	1. Activated HRP conjugates solution 2. HRP labeling solution 3. HRP quencher	3.5-4h	30ug/￥$270 100ug/$275 300ug/$330 1mg/$555 5mg/$1887
ThermoFisher	Samples contain free amino group	1. Activated HRP conjugates solution 2. HRP labeling solution 3. HRP quencher 4. Protein A/G Column	overnight+＞2h	300µg/$652

* How does Abbkine HRP coupling kit remove free HRP (molecular weight 40KD) from binding antibody?

Abbkine HRP coupling kit was designed with this in mind. The kit can provide a low level of free HRP at the end of the reaction. Therefore, no filtration step is required, and the termination solution (quencher) provided in the kit will block the reactive groups of free HRP.

* Protein A/G column in Thermofisher brand is used to remove free HRP.

• Green fluorescence Coupling Kits 

Brand	Sample types	Contents of the kit	Duration of experiment	Size/Price
Abbkine	Samples contain free amino group	1. Activated Green fluorescence conjugates solution 2. labeling solution 3. Purification Column	1.5h	3*20ug/$120 100ug/$140 3*100ug/$160 1mg/$250
Abcam	Samples contain free amino group	1. Activated Green fluorescence conjugates solution 2. labeling solution 3. quencher	0.5-1h	30ug/$265 100ug/$266 300ug/$415 1mg/$645
ThermoFisher	Samples contain free amino group	1. Activated Green fluorescence conjugates solution 2. labeling solution 3. Purification Column	1.5-2h	100µg/$538

* The purpose of the purification column is to separate the unconjugated dye from the coupled product, because the dye is small molecules (molecular weight is about 1KD), and the purification column volume provided by Abbkine is 50KDa, 0.5 ml. The purification column provided by ThermoFisher is also aimed at the purification of antibody (the molecular weight of antibody is about 160KD). If the molecular weight of the protein purified by customers is small or large, they need to equip themselves with appropriate ultrafiltration tubes.

The following recommendations are made for the allocation of ultrafiltration tubes:

The molecular weight of the sample	Ultrafiltration tubes
6<MW<20K	3K
20<MW<60K	10K
60<MW<100K	30K
100<MW<200K	50K
200K<MW	100K

To sum up, which brand of coupling kit would you choose? As the fans of Abbkine, I'd like to summarize the characteristics and advantages of Abbkine LinKine™ products:

• Optimized Scheme-Follow the standard experimental procedure and obtain the best activity or fluorescence with excellent pre-activated dye, protein ratio and patented solution formula.
• The operation is simple and convenient-the operation can be realized in three steps, and the coupling group is bound to a primary amine site of an antibody or other protein.
• Customization-changing molar ratio, reaction buffer, pH value and other parameters can realize different levels of coupling and activity.
• High Efficiency Purification-The kit includes a purification column to ensure rapid and effective removal of unreacted dyes and excellent protein/antibody recovery.