Cell Separations

[ksmith]

When performing cell separations, we currently use microhematocrit centrifugation. We have an old metal 3 sided utility file (non-sharp) to aid in cutting the capillary tubes to recover the retics. We are looking for something to replace this file.

For those who perform cell separations by this method, what tool do you use to aid in this process?

Thanks.

[RR1]

Could the manufacturer recommend a suitable tool?

[AMcCord]

We use a little blade (not sharp) that is about 2-3 inches long and has an edge like a file. Kind a like a tiny hack saw blade. I have no idea in the world what it is or where it came from but you could look over the right section in the hardware store and I'll bet you'd find something similar.

[marquij@caribe.net]

We also do cell separation by microhematocrit centrifugation, but we use plastic tubes for safety which are easy to cut using regular scissors.

[Malcolm Needs ☆]

Many, many years ago (far too many for comfort), when I was briefly working in pure haematology (and not enjoying a moment of it!) we used to use an industrially produced diamond on the end of a pencil like thing for etching names and numbers on slides.

I don't know if these are still available, but if they are, these could possibly do the job.

:confused::confused:

[Dr. Pepper]

I concur with nvelazquez: plastic tubes and a pair of utility scissors make the job easy. We used to use a file and glass tubes but it was sloppy and more dangerous.

[Mabel Adams]

We no longer have the old traditional style of microhematocrit centrifuge.  Our current one only needs to spin Hcts for 2 or 3 minutes.  This means its G force is different from the ones listed in all of the procedures for reticulocyte separation.  Does anyone know what the G force of the old ones was so we can figure out how many minutes to spin in the new ones.

 

Also, the new centrifuge spins at a fixed slant.  Will this alter how the retics rise to the top?

 

Now to go see if I still have my little 40 year old saw.

[SMILLER]

I chuckled as I read some of these posts.  Its funny how facilities have the same little problems that you never think of others having!

 

For years we had a little blade with a serrated edge that worked great for cutting microHct tubes.  Never found out where it came from.  Looked like something that fit inside of a small craft knife handle. Then we lost it.  Looked on-line, asked at hobby stores, never could replace it.  Regular files, even the slimmest, are too fat to do the delicate work without smashing the tube.

 

We have found that sawing with the edge of a scope slide workes pretty well on glass tubes.  Just need to be carefull with it.  Once glass is scored, it will break apart neatly.

 

Scott

[Mabel Adams]

Those little saws used to come in every package of the micropipets that we used with, what was it, our sucker tubes I think.  I can't find mine but my husband has some tiny files called Diamond Needle File Set that I am going to try.  The saw blade from an exacto knife might work too. 

 

I was so hoping you were answering my G force question when I saw a new post on this. 

[cbaldwin]

Those little saws used to come in every package of the micropipets that we used with, what was it, our sucker tubes I think.  I can't find mine but my husband has some tiny files called Diamond Needle File Set that I am going to try.  The saw blade from an exacto knife might work too. 

 

I was so hoping you were answering my G force question when I saw a new post on this. 

.

Try this: RCF (in G forces)= 1.118 x10^-5 x RPM^2 x radius (in cm)

[Mabel Adams]

Thanks, Catherine, but what I need to know is what the G force was of the microhematocrit centrifuges that they used to define the cell separation procedure so I can make sure that my current centrifuges are accomplishing the same force.  I don't know what their radius was, nor what RPM they ran at.  I have found some similar ones on line so I can probably make a reasonable guess.

[Dr. Pepper]

I had a chuckle as well. I saw the little pepper man and thought who's using my avatar? Oh. That was me 3 years ago.

 

So I asked around the (other) elders of the lab. Only one besides me, a hematologist, remembered seeing the little saw blades, about an inch long and shaped like a comb, but couldn't remember where they came from or what they were used for.  My guess had been to score glass tubing before snapping it to make your own pasteur pipets (which I did 35 years ago in my Peace Corps days in West Africa. I don't remember how we snapped the tubing but I do remember heating it in a burner until it softened then pulling the ends apart like taffy to make the thin tips). Thank you Mabel, for remembering the source, but why would anyone besides silly blood bankers want to cut crit tubes? We still go with the plastic-coated safety tubes and a pair of utility scissors.

 

Perhaps someone can help me with a bit of theory about how the retic separation works. I know the patient's retics are less dense and will gather in one end of the tube, but what about the donor retics? Wouldn't they do the same thing? Does something happen to "newly formed" (to quote the Tech Manual) donor RBCs, during storage perhaps, to make them denser?

[SMILLER]

Actually, we use our antique microHct fuge to spin tubes in order to make buffy coat smears in Hematology.  And for these, you want to make a very precise cut.  There is no way you can snip a mylar-wrapped tube and get good results -- it just makes a mess.  We have a vial of glass tubes just for this purpose.

 

Scott