Technical Information

ReadiLink Antibody Conjugation Kits

1. What are the advantages of ReadiLink Antibody Conjugation Kits?
2. What can I label with ReadiLink Antibody Conjugation Kits?
3. What experimental conditions are required?
4. How long does antibody conjugation take?
5. Do I need an extra step to separate out my labeled antibody?
6. How should I store my conjugated antibodies?
7. Still have unanswered questions?

1) What are the advantages of ReadiLink Antibody Labeling Kits?
 

Fast and simple procedure, without further processing

ReadiLink Kits offer a simple antibody solution for labeling microscale amounts (50-100 µg) of antibody for flow cytometry and cell sorting applications in just over an hour. For ease of use each kit contains 2 vials of dye, each vial is for 50 µg of antibody, or other protein greater than 10 kDa in size.

The ReadiLink Antibody Labeling Kits quickly conjugate in two easy steps, label and quench. After the conjugation reaction and addition of quencher buffer, any unbound dye will bind to the quencher and become non-fluorescent.

There are 12 fluorophores available, most of which are ideal for flow cytometry. A wide spectrum of wavelengths ranging from violet to infrared is offered for greater flexibility in multicolor assay design.

For larger scale reactions and conjugations with alternative labels such as enzymes and biotin, please see our range of LYNX Rapid Conjugation Kits^®.

2) What can I label with ReadiLink Antibody Conjugation Kits?
 

The reactive dyes in ReadiLink Antibody Conjugation Kits selectively bind the primary amines of proteins to form a stable carboxamine bond. The kits have been developed to label purified monoclonal or polyclonal antibodies, however other proteins greater than 10 kDa in size can also be labeled. Each kit can label up to 100 µg of antibody (2 x 50 µg reactions).

3) What experimental conditions are required?
 

Antibody preparation for labeling

The antibody should be purified, as all amine containing proteins will be labeled. Antibodies in formats such as ascites, tissue culture supernatant or serum are unsuitable, as well as those containing BSA or gelatin, as all proteins will be conjugated to the fluorophore. Conjugation efficiency is significantly reduced if antibody concentration is less than 1 mg/ml. For optimal labeling efficiency, a concentration of 1-2 mg/ml is recommended.

Buffers and additives for antibody labeling

Antibodies for conjugation should preferably be prepared in phosphate buffered saline (PBS) pH 7.2 -7.4, but HEPES, MES, MOPS or other amine-free buffers are also suitable. Amine containing buffers such as Tris buffer and glycine buffer are unsuitable. If the antibody is in these buffers it will require dialysis against PBS to remove free amines or ammonium salts.

Buffer containing less than 0.1% sodium azide or 10% methanol should not interfere with labeling.

4) How long does antibody conjugation take?
 

Actual hands-on time

It takes just 1-2 min of your time to conjugate an antibody with the ReadiLink Conjugation Kits.

Total conjugation time

The antibody conjugation is complete in 70 min with only two hands-on steps required.

5) Do I need an extra step to separate out my labeled antibody?
 

Antibodies conjugated using the ReadiLink Kit require no post-conjugation clean up processes (column purification, dialysis/desalting). After conjugation and addition of the quencher buffer, any unbound dye will bind to the quencher and become non-fluorescent. The antibody is ready to use immediately.

6) How should I store my conjugated antibodies?
 

The conjugated antibody should be stored at a concentration of at least 0.5 mg/ml. Addition of antimicrobial agents, preservatives, or stabilizers (e.g sodium azide, BSA, glycerol, etc) is strongly recommended for storage at 4°C. For long term storage, the protein conjugates can be lyophilized or divided into single-use aliquots and stored at ≤ –20 °C.

7) Still have unanswered questions?
 

If you have any other questions, contact our technical support team.