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-- - From Margaret Cooley, St. Vincent's Hospital, Darlinghurst Australia...We have tried numerous magnetic separation methods, including Dynabeads, Amerlex-M beads, Biomag and MACS, mostly for human peripheral blood lymphocyte subsets. In our hands, MACS, while having a high startup cost, gives the best results, is gentlest on our fragile cryopreserved cells, and ends up cheapest in the long run. Separating CD4 or CD8 pos cells using directly conjugated beads, we can get 95-99% purity: it may be necessary to manipulate the conditions of flow etc to achieve this with other populations or with indirect beads. We have not observed any effects of the beads on subsequent functions such as proliferation and cytokine production over that seen with antibody alone. The MACS has the advantage over all the other methods that we've tried in that the beads are small enough not to affect scatter on the flow cytometer so subsequent analysis is easy. However, beware clumps, especially when using indirect antibody-bead combinations.
With respect to separating nucleic acids, one of the companies, I think Dynal, has beads designed for this purpose which are commercially available.
-- From: mchugh@jekyll.ucsf.eduWe have used the MACS beads and the Dynabeads; along with polystyrene beads. TheMACS beads have in our hands performed better than the others. The small size does not seem to limit their effectiveness and we have never removed the particles after purification. The purity rates seem high and the cells seem to function and phenotype as you would expect after separation.
-- From: Andreas RadbruchWet greetings from Cologne,
We have quite some experience with MACS, esp. for pre-enrichment for FACS. Due to the small size of the beads they do not interfere with flow cytometry, preety convenient! We have mainly sorted Lymphocytes, stem cells and mega- karyocytes and did not find any detectable interference with cell function. The magnetic fields are not strong by themselves ( below 1 Tesla ) but the field gradient is, making the cells attaching to the ferromagnetic matrix. Some of our work on combined MACS/FACS for stem cell sorting has been published in Cytometry 14, 384-392 (1993). Good luck with your grant!
-- From: Burt Houtz, BD Immuncytometry Systems Wet Greetings from California!In response to your inquiry on MACS, and as a former support representative for this product ( We no longer distribute MACS) - my experience has been quite positive with MACS. I have used it for sorting human CD3, CD4 and CD19 from PBMC (>95% purity), and sorting CD71 positive reticulocytes from whole blood( 70% purity). Also, Thy1.2 positive mouse spleen cells produced about 90% purity. The purity and recovery are largely due to technique improvements and speed. I am not aware of lymphocyte activation from customers I have spoken with, and I have not tested functionality of the sorted cells myself. The "superparamagnetic" properties of the beads prevent magnetic moments existing in the particles away from the magnet, so there should not be any effects on the cells after they have been separated. This works very well as a pre-enrichment tool for subsequent multi-parameter sorting with a FACStar Plus or FACS Vantage.
I am not aware of applications tested on the use of beads for nucleic acid separation.
If you need more information on MACS, call 1-800-FOR-MACS.
-- VIA TELEPHONE FROM: Woody Woodward, Miltenyl salesperson,Woody states that no one has reported any apparent side effect of the magnetic field on the cells or activation of the cells by bead binding. He also states that the company is working on applications involving nucleic acids, but would give no specific details.
--Thank you for the responses. I decided to go with the request for the MACS system.
Kristi Harkins (kharkins@iastate.edu)
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