This product is manufactured by Expedeon, an Abcam company, and was previously called Lightning-Link® HRP Labeling Kit. 701-0004 is the same as the 5 mg size. 701-0003 is the same as the 5 x 1 mg size. 701-0000 is the same as the 3 x 100 ug size. 701-0030 is the same as the 3 x 10 ug size. 701-0010 is the same as the 100 µg size. 701-0002 is the same as the 1 mg size.
Amount and volume of antibody for conjugation to HRP
|
Kit size
|
Recommended
amount of antibody1
|
Maximum
amount of antibody
|
Maximum antibody
volume2
|
|
3 x 10 µg
|
3 x 10 µg
|
3 x 40 µg
|
3 x 10 µL
|
|
100 µg
|
1 x 100 µg
|
1 x 400 µg
|
1 x 100 µL
|
|
3 x 100 µg
|
3 x 100 µg
|
3 x 400 µg
|
3 x 100 µL
|
|
1 mg
|
1 x 1 mg
|
1 x 4 mg
|
1 x 1 mL
|
|
5 x 1 mg
|
5 x 1 mg
|
5 x 4 mg
|
5 x 1 mL
|
|
5 mg
|
1 x 5 mg
|
1 x 20 mg
|
1 x 5 mL
|
1 Recommended amount of antibody will give an HRP:antibody molar ratio of 4:1. Antibodies often also perform well at the maximum amount of antibody which is 1:1 molar ratio.
2 Ideal antibody concentration is 1mg/ml. 0.5 - 1 mg/ml can be used if the maximum antibody volume is not exceeded. Antibodies > 4 mg/ml or < 0.5 mg/ml should be diluted /concentrated.
Buffer Requirements for Conjugation
Buffer should be pH 6.5-8.5.
Buffer constituent compatibility
If a concentration is shown, then the constituent should be no more than the concentration shown. If several constituents are close to the limit of acceptable concentration, then this can inhibit conjugation.
|
Compatible buffer constituents
|
|
50mM / 0.6% Tris1
|
0.1% BSA
|
50% glycerol
|
|
0.1% sodium azide2
|
PBS
|
Potassium phosphate
|
|
Sodium chloride
|
HEPES
|
Sucrose
|
|
Sodium citrate
|
EDTA
|
Trehalose
|
1 Tris buffered saline is almost always ≤ 50 mM / 0.6%
2 Sodium azide irreversibly inhibits HRP; antibodies with azide should be purified before using the HRP conjugation kit.
|
Incompatible buffer constituents
|
|
Thiomerosal
|
Proclin
|
Glycine
|
|
Arginine
|
Glutathione
|
DTT
|
If a constituent of the buffer containing your antibody or protein is not listed above, please check the FAQ or contact us.
Only purified antibodies are suitable for use, ie. where other proteins, peptides, or amino acids are not present: antibodies in ascites fluid, serum or hybridoma culture media are incompatible.
Storing and handling conjugation kits
Lyophilized Lightning-Link® components are hygroscopic.
Kits are intentionally shipped at ambient temperature with silica gel to avoid exposure to moisture. Upon receipt, store the kit frozen and protect from moisture. Before opening the outer container, allow the lyophilized components to reach room temperature to minimize condensation.
