Non-mammalian and non-bacterial based / Cell detachment solution
Accumax is a ready to use non-mammalian, non-bacterial replacement for all applications of trypsin and collagenase in tissue dissociation, cell counting, and dissolving cell clumps such as spheroids. Accumax contains the same enzymes as Accutase and is a direct replacement for collagenase, and can be used to increase the accuracy of a cell count. It can also be added to a clumpy sample on a cell sorter to extend the sort time of the sample.
>> Click here to Accutase™ Cell Dissociation Solution
A few cell lines that Accumax has been shown to detach without harm:
|vascular endothelial cells
|vascular smooth muscle cells
|primary chick embryo neuronal cells
|bone marrow stem cells
|adherent CHO cells
|adherent BHK cells
|testicular germ cells
|M24 and A375 metastatic melanoma
|gliomas U251 and D54
|HT1080 fibrosarcoma cells
|Sf9 insect cells
Accumax performs exceptionally well for:
Accumax is used to digest primary tissue, create single cell suspensions for cell counts, and declump cells for magnetic or flow cytometry cell sorting and removing from artificial growing matrixes.
|virus growth assay
|analysis of cell surface markers
|tumor cell migration assays
|routine cell passage
|quiescence assays by serum starvation
|transformation assays by oncogenetransfection
|transfectionneural crest cell migration assays
|production scale-up (bioreactor)
|neural crest cell migration assays
|Various constructs of genetically engineered CHO cells, BHK cells, and a hybridoma were grown in suspension in serum-free or protein-free medium.
Representative cell aliquots were treated with an equal volume of PBS or Accumax and incubated for 5 minutes at 37C. Cell number was then determined with a Coulter Counter.
This protocol for using Accumax to dissociate cells from primary tissue is a general-purpose protocol and may not be applicable to all tissue types. The individual investigator needs to optimize the conditions for his/her tissue specimens. Keep in mind that Accumax is a powerful enzyme mixture that can potentially dissolve not only the connective tissue of solid tissue but some fragile cell types as well if not closely monitored.
Accumax (Should be defrosted overnight in the refrigerator or in a bucket of room temperature water - not a 37°C bath)
DPBS (calcium and magnesium free)
Culture medium, i.e., DMEM/F12 with 10 – 20% FBS (or other appropriate media)
Pipettes - 1 ml, 10 ml
Petri dishes -100 mm, non-tissue culture grade
T25 culture flasks
Centrifuge tubes, 15-50 ml, depending upon the amount of tissue being processed
If cell isolation is from a soft tissue (such as liver):
Regardless of what the manufacturers of cell counters tell you, when counting cells with either a manual or automated method, the accuracy and the reproducibily of the counts will be increased if the cells are not clumped together. Accumax solution can be used to dissociate clumpy cells that are being counted. Accumax is gentle enough that an aliquot of it can be added to an aliquot of cells and allowed to set for a period of time without damaging the cells.