Re[2]: [DIYbio] An update on my effort to isolate viable adipose derived stem cells for therapeutic reinjection and a call for advice

thanks for the feedback. very useful. So from your reading, these
processes end with the multicell SVF which is what they use as is in
their applications? i would consider using the filter at the end which
some of the protocols use. D you understand that to be for isolating he
ADSC's or for purifying all the live cells in the svf from surrounding
material?
Regarding contamination with fungi and the like, that's very important.
there are some protocols where the material remains in the same syringe
keeping it sterile.



------ Original Message ------
From: "Dakota Hamill" <dkotes@gmail.com>
To: "diybio@googlegroups.com" <diybio@googlegroups.com>
Sent: 12/21/2020 2:14:44 PM
Subject: Re: [DIYbio] An update on my effort to isolate viable adipose
derived stem cells for therapeutic reinjection and a call for advice

>Pretty interesting pictures in some of the articles. Amazing you can
>take out adipose tissue, diggest it, spin it, and harvest a plethora
>of cell types.
>
>As you mentioned, it looks like only 5% of that final fraction is
>adipose derived stem cells. I imagine it will be difficult to select
>only the ADSC's from that fraction with low-budget equipment. A flow
>cytometry machine would be able to do it. A very careful hand and a
>tiny pulled glass syringe might under a microscope.
>
>I've done 0 human cell line culture in my life but even the lab next
>to us who has an ozonator, secure room, hoods, everything, often
>complains of contamination of fungal spores etc. It's not easy!
>
>You may just take the cell mixture found in the VSF and culture that
>and see what cells tend to dominate in it. It may be the stem cells
>do well in culture and out-grow other types. It may be they do not,
>and further purification to a monoculture is needed. How you'd do
>that, I have no idea but it's interesting.
>
>On Fri, Dec 18, 2020 at 11:10 AM Frank Garcia <fgarcia0007@gmail.com> wrote:
>>
>>
>> So I've scoured the internet for papers and guides to nonenzymatic procedures that can be done with simple inexpensive equipment and which yields an usable quantity of fat stem cells that are alive and well.
>>
>> I've read a few and must say I'm not entirely sure if I understand the required steps.
>> Each of the following describes variations of the procedure. Almost all add some fancy element in one of the steps like a commercial kit, specialized syringe with filters built in , or custom made contraptions - get a load of this "Custom-made metallic disarmable pistons, concave cell-adhesive gaskets, closed cubic unit harnessing 3 different-sized sets of blade grids on each luer-lock port, rotating canal at the center of the cube to control the flow of the lipoaspirate, http://links.lww.com/PRSGO/B308).".
>>
>>https://www.nature.com/articles/s41598-017-10710-6#Tab3
>>https://journals.lww.com/prsgo/fulltext/2020/02000/a_3_step_mechanical_digestion_method_to_harvest.27.aspx
>>https://stemcells-aesthetica.com/isolation-process-of-stem-cells-from-fatty-tissue/
>>http://www.irvinesci.com/protocol-for-mesenchymal-stem-cell-isolation
>>
>> I'd like to use these to put together a protocol that uses no expensive or custom made equipment. I'd like to know if the plating and incubation is necessary and is filtering necessary? Also, is the VSF the end point or can ADSCs be isolated from VSF? any other feedback welcome.
>>
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