Small Animal In Vivo Imaging Antibody Labeling
Antibodies that are fluorescently labeled for use in animals have more stringent requirements than antibodies labeled for other types of applications. The SAIVI™ Rapid Antibody Labeling Kits are uniquely designed to provide the rigorous requirements to control the degree of labeling (DOL). This control is accomplished by limiting the number of fluorescent molecules covalently attached to the antibody (Figure 1). It is crucial to restrict the DOL for antibodies used in vivo compared to other types of antibody applications, in order to maximize the desired signal at its target location while minimizing accumulation in the liver (Figure 2). Antibodies labeled with the SAIVI™ Rapid Antibody Labeling Kit are ideal for in vivo use.
Non-invasive optical imaging in small animals is best achieved with antibodies or proteins labeled with near-infrared (NIR) fluorophores. AlexaFluor® dyes that emit in the NIR 700–900 nm spectrum are ideal for labeling antibodies to be used for in vivo imaging. Dyes in this range are critical to maximize the penetration of the signal while reducing autofluoresence from nontarget tissues. This highly sensitive detection can often be viewed to 3 mm or more, depending on the instrument used for visualization.
Figure 1. Conjugated antibodies used for in vivo applications have a limited and controlled number of labels attached.
Figure 2. An anti-CEA antibody targeting colon cancer xenografts in mice show the optimal DOL for the fluorescence antibody to be 2.4, while limiting intensity in the liver, as imaged with the CRi Maestro™ In Vivo Imaging System.
Get the best results with antibodies for in vivo imaging applications
In addition to the DOL control reagent, the SAIVI™ Rapid Antibody Labeling Kits include a purification column with an easy-to-use pre-measured slurry that is specifically designed for a fast, 10 minute prep time and short wash step. The removal of azide is not required as it is for other columns, offering additional time savings. This protocol reliably results in high yields of ready-to-use antibody. The table below compares the use of these kits, and shows multiple advantages of using the SAIVI™ Rapid Antibody Labeling Kits for all in vivo applications.
Rapid SAIVI™ Antibody Labeling Kits vs. other standard antibody labeling kits
|Standard kits||Rapid SAIVI™ kit|
|DOL Control Reagent||None, kits optimized for DOL in 4-7 range not in vivo applications||Optimal 1.5-3 DOL for in vivo applications|
|Time Required for purification||30 min||<10 min|
|Animal Readiness post-column purification||Sterile filtration; azide removal required||Ready to inject|
Need to label other antibody amounts?
|IgG amount||Attachment of label||Products|
|<1–20 µg||Site-specific||Zenon® Antibody Labeling Kits|
|<100 µg||Site-specific||SiteClick™ Antibody Labeling Kits|
|10–20 µg||Conventional covalent||APEX® Antibody Labeling Kits|
|20–100 µg||Conventional covalent||Microscale Protein Labeling Kits|
|100 µg||Conventional covalent||Antibody Labeling Kits|
|1 mg||Conventional covalent||Protein Labeling Kits|
|0.5–3 mg||Conventional covalent||SAIVI™ Rapid Antibody Labeling Kits|