A Technique to Assess the In Vivo Localization of Tumor-Specific Antibodies
Published: November 30, 2023
Abstract
Source: Shivange, G., et al. Analyzing Tumor and Tissue Distribution of Target Antigen Specific Therapeutic Antibody. J. Vis. Exp. (2020).
This video demonstrates a technique to study the in vivo localization of tumor-specific antibodies in a mouse tumor xenograft model. Upon subcutaneously injecting cancer cells into mice for tumor formation, fluorescently-labeled antibodies are injected that bind to tumor-specific receptors, enabling in vivo assessment of antibody localization via imaging.
Protocol
All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Expression and purification of antibodies Maintenance of Chinese hamster ovary (CHO) cells Grow CHO cells in FreeStyle CHO Media supplemented with commercially available 1x glutamine supplement at 37 ˚C shaking at 130 rpm with 5% CO2 using delong Erlenmeyer flasks either glass or disposable.<…
開示
The authors have nothing to disclose.
Materials
| FreeStyle CHO media | Gibco Life Technologies | Cat # 12651-014 | |
| Anti-Anti (100X) | Gibco Life Technologies | Cat # 15240-062 | |
| Anti-Clumping Agent | Gibco Life Technologies | Cat # 01-0057DG | |
| BD Insulin Syringe | BD BioSciences | Cat #329420 | |
| Caliper IVIS Spectrum | PerkinElmer | Cat #124262 | |
| CHO CD EfficientFeed B | Gibco Life Technologies | Cat #A10240-01 | |
| Corning 500 mL DMEM (Dulbecco's Modified Eagle's Medium) | Corning | Cat # 10-13-CV | |
| Corning 500 mL RPMI 1640 | Corning | Cat # 10-040-CV | |
| Cy5 conjugated Anti-Human IgG (H+L) | Jackson ImmunoResearch | Cat # 709-175-149 | |
| GlutaMax-I (100X) | Gibco Life Technologies | Cat # 35050-061 | |
| HiPure Plasmid Maxiprep kit | Invitrogen | Cat # K21007 | |
| HiTrap MabSelect SuRe Column | GE Healthcare | Cat # 11-0034-93 | |
| Infusion | Takara BioScience | STO344 | |
| IRDye 800CW NHS Ester | LI-COR | Cat # 929-70020 | |
| Isoflurane, USP | Covetrus | Cat # 11695-6777-2 | |
| Lubricant Eye Ointment | Refresh Lacri-Lube | Cat #4089 | |
| Matrigel | Corning | Cat # 354234 | |
| PEI transfection reagent | Thermo Fisher | Cat # BMS1003A | |
| Slide-A-Lyzer Dialysis Cassettes | Thermo Scientific | Cat # 66333 | |
| Steritop Vacuum Filters | Millipore Express | Cat #S2GPT02RE | |
| Trypsin-EDTA | Gibco Life Technologies | Cat # 15400-054 | |
| Experimental Models: Cell lines | |||
| Human: OVCAR-3 | American Type Culture Collection | ATCC HTB-161 | |
| Human: CHO-K cells | Stable transformed in our lab | ATCC CCL-61 | |
| Mouse: 4T1 | Kind gift from Dr. Chip Landen, UVA | ||
| Mouse: MC38 | Kind gift from Dr. Suzanne Ostrand-Rosenberg, UMBC | Authenticated by STR profiling | |
| Mouse: MC38 hFOLR1 | Generated in our laboratory (This paper) | ||
| Experimental Models: Animal | |||
| Mice: athymic Nude Foxn1nu/Foxn1+ | Envigo | Multiple Orders | |
| Mice: NOD.Cg Prkdcscid Il2rgtm1Wjl/SzJ | Jackson Laboratory | Multiple Orders |
