[Histonet] Manipulation time of human muscle biopsies and formation of freezing artifacts.

[Mauricio S. Morais]

Hello Histonet community.

Because I'm new into the histology "word" I've been facing some
unexpected (to me) challenges in my actual Laboratory work.
My background is originally from Organic and Analytical Chemistry,
Teaching, and a bit of Biochemistry (my MSc degree).
I'm kind of new in the country too (been here less than 2 years) and
English is not my primary language, so please accept my apologies for
any errors within this text.

I'm assign to reorganize and analyze human muscle biopsies, stored in
boxes distributed among 4 racks inside a Cryoplus cryofreezer.
Seeking for information on how to work safely and at the same time not
compromise the integrity of the stored samples, I found the Histonet
Histosearch archives.
They were very helpful providing answer about most of the common issues
related with formation of freezing artifacts and possible solutions to
the problem at different stages the samples can generate it, from the
biopsy to the cryostat slice machine.

Because of the enormous amount of samples that'll need to be handle at
one time during the reorganization of our cryofreezer, an issue about
"time"  became critical (in my opinion) raising some questions:

1-How long can a box (full of biopsies), or the hole rack be kept out of
the cryofreezer, while I'm updating data and re-arranging boxes between
racks, without compromise the biopsies and generate (and/or enhancing
the chances of creating) freezing artifacts?

2-Does any damage may occur if I have several biopsies floating inside a
dewar container while I'm making notes or grouping them into other dewar
containers to a new order? 

A website information (from www.asymptote.co.uk) says that tissue inside
straws immersed in liquid Nitrogen when exposed to air at room
temperature have their temperature raised to -50C in 40 seconds. Thawing
may start around -10C or early.

I was told they (racks, boxes) could be out for about 5 minutes before
some damage can start to affect the biopsies inside it. None of them
been hold by hands during all this time, but on the bench top (boxes) or
floor (racks) while the work is done.
I was also told that 5 minutes is away too long...

For me it seems impossible to safely remove a rack (containing a dozen
boxes) from the cryofreezer, make notes or, for example, remove one box
from the rack to add a new sample and return it to the cryofreezer in 40
seconds. Reminding that I'm working with cryogloves over my lab.
gloves... I was never able to open a box without remove the cryogloves
first.
Maybe I need more training...

I would greatly appreciate any comments from this expert community to
help me solve my immediate problems (above) and improve my knowledge and
safety at work place.
 

Thank you all.


Mauricio S. Morais
Senior Research Technician
Nutrition Exercise Physiology and Sarcopenia Laboratory (NEPS)
TUFTS University
Jean Mayer USDA Human Nutrition Research Center on Aging
711 Washington Street, Rm 436
Boston, MA 02111
(617) 556-3226
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