There have been some studies that have prestained cells with submicron fluorescent microspheres, which are generally inert, highly persistent (months for tissues) and intensely fluorescent before transplantation. Follow this link http://www.probes.com/handbook/sections/1406.html and look for the topic: Transplantation and Migration Studies with Fluorescent Microspheres for some leading references. This is also a good way to mark transplanted tissues to look for regrowth and acceptance. The microspheres can potentially be sterilized by ozone or by heating below their softening point or maybe even ultrafiltration of the very small microspheres (<0.1 micron). Although we have not tried them for this purpose, I recommend trying 0.02 um yellow-green fluorescent azide-free microspheres for this purpose. http://www.probes.com/servlets/product?region=USA&item=8795 The same methods MAY be useful for tracing other cells that have been injected into an organism. This method seems highly underutilized for cell tracing but is inexpensive and I believe will prove to be nontoxic and very sensitive. An additional advantage is that we make microspheres that can be excited at 488 nm with five different emission maxima far removed from 488 nm (at about 560, 605, 645, 685 and 720 nm) http://www.probes.com/handbook/figures/0604.html that would allow the FL 1 (and even FL2 and FL 3 channels) to be used for additional detection reagents or one could potentially stain more than one type of cell with different colors of microspheres and trace these separately or stain the same cells that are to be injected at different times and trace these. The microspheres probably attach themselves the the cell's surfaces by hydrophobic forces or may be injested. Thus, the surface charge may have some effect on the adsorption and retention. The studies I have looked at indicate that the binding appears to be irreversible, meaning that once attached the microspheres do not transfer between cell populations. One could use a dead cell stain such as propidium iodide to look for viability of the cells that have been labeled. "Otten, Gillis" wrote: > A colleague is transplanting human cells into mice and would like to be able > to identify, using immunofluorescence and flow cytometry, the transplanted > human cells in tissues isolated from the mice. Can anyone recommend > monoclonal antibody conjugates that will stain human, but not mouse cells? > The human cells are not of hematopoietic origin. Also, because cell surface > markers are unlikely to survive the cell isolation procedure, we would > prefer an anti-human antibody that reacts with an intracellular antigen, > although we wouldn't rule out testing mAb that bind cell surface markers. > Any suggestions or comments based on experience with human cell-into-mouse > transplants would be much appreciated. Thanks. > > Gib Otten > Chiron Corp. > Emeryville, CA 94608 > mailto:Gillis_Otten@chiron.com > > ** ** ATTENTION! This message contains information (including any appended information, > such as copied or forwarded messages and attachments, and any message to which this is > appended) which may be confidential and/or privileged. Unless you are the addressee (or > authorized to receive for the addressee), you may not use, copy or disclose to anyone the > message or any information contained in the message. If you have received the message > in error, please advise the sender by reply e-mail, and delete the message. Thank you > for your assistance. ** **
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