[Histonet] RE: Frozen sections for Immunoflouresence

[Tony Henwood (SCHN)]

Hi Jim,

You mention that the OCT seems a little softer.
Could it be that the sections are thicker than you usually produce. Check the 
Frozen section H&E to check this.
If sections are thicker they will tend to detach from the slides.

Regards 
Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC), FFSc(RCPA) 
Laboratory Manager & Senior Scientist, the Children's Hospital at Westmead
Adjunct Fellow, School of Medicine, University of Western Sydney 
Tel: 612 9845 3306 
Fax: 612 9845 3318 
Pathology Department
the children's hospital at westmead
Cnr Hawkesbury Road and Hainsworth Street, Westmead
Locked Bag 4001, Westmead NSW 2145, AUSTRALIA 

-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu 
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Vickroy, Jim
Sent: Wednesday, 3 September 2014 12:51 AM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Frozen sections for Immunoflouresence

Lately we are experiencing an issue where the tissue sections are coming off of 
the slide during the staining process.   We have always used positive charged 
slides and have not experienced this problem on a routine case.   We do not fix 
the slides but allow them to air dry for Immunology.  Some have suggested a 
quick fixation in acetone or 95% ETOH.   This doesn't seem to correct the 
problem and we don't find the IF stains work as well.   We have tried a 
different batch of slides and are looking into a different kind of coating for 
the slides.   Our regular frozen sections stay on the slides.  We are using 
OCT.   The only other thing I have noticed is that maybe it's my imagination 
but the OCT in the block surrounding the kidney biopsy seems a little softer 
than normal.   I did try a different mounting medium and maybe the softness was 
my imagination.   Has anyone else experienced this problem and if so how did 
you correct it?   The lab is air conditioned so I don't think the humidity is 
the main factor.  The Immunology staff changed all of their reagents so we have 
reduced that variable hopefully.   We are using Leica positive charged slides.  
(I have found that the Apex control box slides from Leica are a little better 
than the regular Apex slides.   Any ideas?

Jim


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[Histonet] RE: IHC Validation

[Algeo, Lacie A]

I have used the CAP published 'Quality Management in Anatomic Pathology' and it 
has been very helpful with this:

"For a well-characterized antibody  with a limited spectrum of antigenic 
targets, like chromogranin, or prostate specific antigen, the validation can be 
limited.  A panel of 10 positive and 10 negative neoplasms would be sufficient 
in this setting.  For an antibody that is not well characterized and/or has a 
wide range of reported activity, a more extensive validation is necessary.  The 
number of tissues tested should in this circumstance be large enough to 
determine whether the staining profile matches that previously described. "

We use cases known to be positive or negative for a given antibody or we 
request tumor types that are known to be positive or negative.  We do 10 of 
each.  If an antibody has a wide range of positivities (i.e. % positive like 
c-Myc or FOXP1), we try to have a range of different positivities for review.

Hope this helps,
Lacie

Lacie Algeo, HTL (ASCP) MBCM
Histology Supervisor
Providence Sacred Heart Medical Center Laboratory
101 W 8th Avenue
L-2
Spokane, WA 99204
509-474-4418
FAX 509-474-2052
lacie.al...@providence.org


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[Histonet] Take me out of the mailing list

[Ferrer, Joselito]

Please take me out of the mail list.

Thank you


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[Histonet] Adhesive coated slides verses positive charged slides

[Vickroy, Jim]

Does anyone have any experience on which slides work better for frozen sections 
and immunofluorescence stains..adhesive coated or positive charged?

Does anyone else find the selection of a proper slide confusing or is it just 
my age?   I understand the basis behind hydrophobic, hydrophilic, adhesive 
coated, and positive charged however we have thought our Leica Apex slides were 
positive charged all along.

Thanks

Jim


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RE: [Histonet] Non-cert. histo classes

[Jennifer MacDonald]

I agree with Tim.  Any additional skills in histology will make them more 
marketable.  EM and paraffin microtomy are very similar in principle.

As of yet an AA/A is not required to take the HT (ASCP) certification 
exam.  Route 1 states "graduation from a NAACLS accredited program" and 
there are still some NAACLS accredited programs that offer a certificate 
and do not require an AA or AS.  The NSH appealed to NAACLS to change one 
of the Standards to require all NAACLS HT programs to either award an 
AA/AS degree or require students to have one before earning the 
certificate.  The changes to the standard went out for public comment at 
the beginning of the summer.  The NAACLS BOD meeting is September 18-19. 
The Board will vote then to either recommend or not recommend the changes 
to the standard.  If it is approved by the BOD there will be a transition 
period to allow those programs that do not offer or require an AA/AS to 
meet the standard.This does not affect those that are already certified, 
nor does it change the ASCP requirements.

Jennifer



From:   "Morken, Timothy" 
To: "'Jon Krupp'" , 
"histonet@lists.utsouthwestern.edu" 
Date:   09/02/2014 10:38 AM
Subject:RE: [Histonet] Non-cert. histo classes
Sent by:histonet-boun...@lists.utsouthwestern.edu



Jon,

It is not a bad idea. Ideally a person going into the field would have a 
good formal education in the field. However, 99.9% of the people working 
in the field did not go through a formal program, but learned on the job. 
Therefore, a person who was exposed to ANY formal education in paraffin 
histotechnology (processing, cutting, staining, special stains) would be 
well ahead- OJT is highly variable in quality as you might guess. A lot of 
what is learned in biological EM is transferable to paraffin - fixation, 
processing, even sectioning principles are the same. The difference is 
really in medium and staining chemicals, and of course, the microscope 
used. 

There are obviously a lot more jobs in histology than in EM. Biotech does 
not necessarily require certification and it is not needed as a regulatory 
requirement of their work (and a combined EM/histotech is more valuable; 
throw in some DNA/RNA work (ISH, FISH, PCR and you have a supertech!). 
Hospitals and other medical labs usually do not require certification 
(like or not!) for entry level (or even higher levels in many cases) but 
if they want you to have it they will usually have a time period that they 
require you to get it - "certification eligible" or "certification within 
one year" or something like that. Many of our histotechs came from the 
UCSF research labs where they learned a bit of paraffin sectioning and 
then applied in our lab. All have done well and all have gone on to get 
certified.

Acquiring certification requires working one year under a board certified 
pathologist, and taking a test. It takes some study, but that is the route 
most people take.

The most important part is that certification now requires an AA degree at 
the minimum with certain levels of biology and chemistry courses. Those at 
Delta would meet that standard pretty easily if they are in the EM program 
anyway. 

Tim Morken
Supervisor, Histology, Electron Microscopy and Neuromuscular Special 
Studies
UC San Francisco Medical Center
San Francisco, CA


-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu [
mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Jon Krupp
Sent: Tuesday, September 02, 2014 9:00 AM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Non-cert. histo classes

Greetings

I am a regular reader of this list, but from a peripheral setting.

I work in an electron microscopy certificate program. Our students are 
prepared to do either or both certificates in EM, biological and 
materials.

During their biological training, students are instructed in thin 
sectioning, specimen prep., etc. I would like to get ideas about how 
valuable adding more light microscopy specimen prep, staining, and 
sectioning might be for these students. Would it be good for them to have 
these skills and knowledge but not a histotech cert.?

I thought about doing a full cert. program, but am daunted by the 
requirements to set one up and I don't want to compete with other better 
established programs near by.

Bottom line is I would like to know if teaching basic histo tech skills, 
w/o certification, is a viable path. Could students leverage these skills 
into jobs at non-health care type facilities? Could having these skill 
help them complete a certificated program if they wanted to take that 
direction later? 

Your input will be valuable to both me and my students.

Thanks

Jon

Jonathan Krupp
Applied Science, Business & Technology
San Joaquin Delta College
5151 Pacific Ave.
Stockton, CA  95207
209-954-5284
jkr...@deltacollege.edu

Find us on Facebook @
Electron Microscopy at SJ Delta College









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Re: [Histonet] Non-cert. histo classes

[Pam Marcum]

I agree with Tim the more you know the better off you are and if you have some 
skills in Histology and can do EM that is a plus.  Any form of molecular is a 
plus now and many companies do not even require certification in any field as 
long as you have the science and background to do the work.  It is a rare thing 
to find someone who can do it all in the microscopy field and I would think a 
good set of skills to be able to present.  
  
EM appears to be in the process of becoming more useful again.  When I learned 
EM years ago it was in a lab with some help from an EM person in another 
department and on my own.  Thank goodness I was too young and dumb to know how 
daunting it could have been to go that route.  Unfortuantely I have not used 
those skills in many years.  
  
Pam Marcum 
UAMS 

- Original Message -

From: "Timothy Morken"  
To: "Jon Krupp" , "Histonet" 
 
Sent: Tuesday, September 2, 2014 12:35:20 PM 
Subject: RE: [Histonet] Non-cert. histo classes 

Jon, 

It is not a bad idea. Ideally a person going into the field would have a good 
formal education in the field. However, 99.9% of the people working in the 
field did not go through a formal program, but learned on the job. Therefore, a 
person who was exposed to ANY formal education in paraffin histotechnology 
(processing, cutting, staining, special stains) would be well ahead- OJT is 
highly variable in quality as you might guess. A lot of what is learned in 
biological EM is transferable to paraffin - fixation, processing, even 
sectioning principles are the same. The difference is really in medium and 
staining chemicals, and of course, the microscope used. 

There are obviously a lot more jobs in histology than in EM. Biotech does not 
necessarily require certification and it is not needed as a regulatory 
requirement of their work (and a combined EM/histotech is more valuable; throw 
in some DNA/RNA work (ISH, FISH, PCR and you have a supertech!). Hospitals and 
other medical labs usually do not require certification (like or not!) for 
entry level (or even higher levels in many cases) but if they want you to have 
it they will usually have a time period that they require you to get it - 
"certification eligible" or "certification within one year" or something like 
that. Many of our histotechs came from the UCSF research labs where they 
learned a bit of paraffin sectioning and then applied in our lab. All have done 
well and all have gone on to get certified. 

Acquiring certification requires working one year under a board certified 
pathologist, and taking a test. It takes some study, but that is the route most 
people take. 

The most important part is that certification now requires an AA degree at the 
minimum with certain levels of biology and chemistry courses. Those at Delta 
would meet that standard pretty easily if they are in the EM program anyway. 

Tim Morken 
Supervisor, Histology, Electron Microscopy and Neuromuscular Special Studies 
UC San Francisco Medical Center 
San Francisco, CA 


-Original Message- 
From: histonet-boun...@lists.utsouthwestern.edu 
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Jon Krupp 
Sent: Tuesday, September 02, 2014 9:00 AM 
To: histonet@lists.utsouthwestern.edu 
Subject: [Histonet] Non-cert. histo classes 

Greetings 

I am a regular reader of this list, but from a peripheral setting. 

I work in an electron microscopy certificate program. Our students are prepared 
to do either or both certificates in EM, biological and materials. 

During their biological training, students are instructed in thin sectioning, 
specimen prep., etc. I would like to get ideas about how valuable adding more 
light microscopy specimen prep, staining, and sectioning might be for these 
students. Would it be good for them to have these skills and knowledge but not 
a histotech cert.? 

I thought about doing a full cert. program, but am daunted by the requirements 
to set one up and I don't want to compete with other better established 
programs near by. 

Bottom line is I would like to know if teaching basic histo tech skills, w/o 
certification, is a viable path. Could students leverage these skills into jobs 
at non-health care type facilities? Could having these skill help them complete 
a certificated program if they wanted to take that direction later? 

Your input will be valuable to both me and my students. 

Thanks 

Jon 

Jonathan Krupp 
Applied Science, Business & Technology 
San Joaquin Delta College 
5151 Pacific Ave. 
Stockton, CA  95207 
209-954-5284 
jkr...@deltacollege.edu 

Find us on Facebook @ 
Electron Microscopy at SJ Delta College 









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RE: [Histonet] Non-cert. histo classes

[Morken, Timothy]

Jon,

It is not a bad idea. Ideally a person going into the field would have a good 
formal education in the field. However, 99.9% of the people working in the 
field did not go through a formal program, but learned on the job. Therefore, a 
person who was exposed to ANY formal education in paraffin histotechnology 
(processing, cutting, staining, special stains) would be well ahead- OJT is 
highly variable in quality as you might guess. A lot of what is learned in 
biological EM is transferable to paraffin - fixation, processing, even 
sectioning principles are the same. The difference is really in medium and 
staining chemicals, and of course, the microscope used. 

There are obviously a lot more jobs in histology than in EM. Biotech does not 
necessarily require certification and it is not needed as a regulatory 
requirement of their work (and a combined EM/histotech is more valuable; throw 
in some DNA/RNA work (ISH, FISH, PCR and you have a supertech!). Hospitals and 
other medical labs usually do not require certification (like or not!) for 
entry level (or even higher levels in many cases) but if they want you to have 
it they will usually have a time period that they require you to get it - 
"certification eligible" or "certification within one year" or something like 
that. Many of our histotechs came from the UCSF research labs where they 
learned a bit of paraffin sectioning and then applied in our lab. All have done 
well and all have gone on to get certified.

Acquiring certification requires working one year under a board certified 
pathologist, and taking a test. It takes some study, but that is the route most 
people take.

The most important part is that certification now requires an AA degree at the 
minimum with certain levels of biology and chemistry courses. Those at Delta 
would meet that standard pretty easily if they are in the EM program anyway. 

Tim Morken
Supervisor, Histology, Electron Microscopy and Neuromuscular Special Studies
UC San Francisco Medical Center
San Francisco, CA


-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu 
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Jon Krupp
Sent: Tuesday, September 02, 2014 9:00 AM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Non-cert. histo classes

Greetings

I am a regular reader of this list, but from a peripheral setting.

I work in an electron microscopy certificate program. Our students are prepared 
to do either or both certificates in EM, biological and materials.

During their biological training, students are instructed in thin sectioning, 
specimen prep., etc. I would like to get ideas about how valuable adding more 
light microscopy specimen prep, staining, and sectioning might be for these 
students. Would it be good for them to have these skills and knowledge but not 
a histotech cert.?

I thought about doing a full cert. program, but am daunted by the requirements 
to set one up and I don't want to compete with other better established 
programs near by.

Bottom line is I would like to know if teaching basic histo tech skills, w/o 
certification, is a viable path. Could students leverage these skills into jobs 
at non-health care type facilities? Could having these skill help them complete 
a certificated program if they wanted to take that direction later? 

Your input will be valuable to both me and my students.

Thanks

Jon

Jonathan Krupp
Applied Science, Business & Technology
San Joaquin Delta College
5151 Pacific Ave.
Stockton, CA  95207
209-954-5284
jkr...@deltacollege.edu

Find us on Facebook @
Electron Microscopy at SJ Delta College









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RE: [Histonet] Non-cert. histo classes

[Nails, Felton]

My opinion is that it is a good idea because many hospitals are shutting down 
their EM department because of a lack of skilled workers.
So the more skills they have the better for them.
Let me know if any of your students want to move to Houston Texas.

Felton Nails
Texas Children's Hospital 
832-824-1248

-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu 
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Jon Krupp
Sent: Tuesday, September 02, 2014 11:00 AM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Non-cert. histo classes

Greetings

I am a regular reader of this list, but from a peripheral setting.

I work in an electron microscopy certificate program. Our students are prepared 
to do either or both certificates in EM, biological and materials.

During their biological training, students are instructed in thin sectioning, 
specimen prep., etc. I would like to get ideas about how valuable adding more 
light microscopy specimen prep, staining, and sectioning might be for these 
students. Would it be good for them to have these skills and knowledge but not 
a histotech cert.?

I thought about doing a full cert. program, but am daunted by the requirements 
to set one up and I don't want to compete with other better established 
programs near by.

Bottom line is I would like to know if teaching basic histo tech skills, w/o 
certification, is a viable path. Could students leverage these skills into jobs 
at non-health care type facilities? Could having these skill help them complete 
a certificated program if they wanted to take that direction later? 

Your input will be valuable to both me and my students.

Thanks

Jon

Jonathan Krupp
Applied Science, Business & Technology
San Joaquin Delta College
5151 Pacific Ave.
Stockton, CA  95207
209-954-5284
jkr...@deltacollege.edu

Find us on Facebook @
Electron Microscopy at SJ Delta College









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[Histonet] Non-cert. histo classes

[Jon Krupp]

Greetings

I am a regular reader of this list, but from a peripheral setting.

I work in an electron microscopy certificate program. Our students are prepared 
to do either or both certificates in EM, biological and materials.

During their biological training, students are instructed in thin sectioning, 
specimen prep., etc. I would like to get ideas about how valuable adding more 
light microscopy specimen prep, staining, and sectioning might be for these 
students. Would it be good for them to have these skills and knowledge but not 
a histotech cert.?

I thought about doing a full cert. program, but am daunted by the requirements 
to set one up and I don't want to compete with other better established 
programs near by.

Bottom line is I would like to know if teaching basic histo tech skills, w/o 
certification, is a viable path. Could students leverage these skills into jobs 
at non-health care type facilities? Could having these skill help them complete 
a certificated program if they wanted to take that direction later? 

Your input will be valuable to both me and my students.

Thanks

Jon

Jonathan Krupp
Applied Science, Business & Technology
San Joaquin Delta College
5151 Pacific Ave.
Stockton, CA  95207
209-954-5284
jkr...@deltacollege.edu

Find us on Facebook @
Electron Microscopy at SJ Delta College









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[Histonet] RE: Frozen sections for Immunoflouresence

[Burnett, Brandy]

Are you using pre-cut control slides?


From: histonet-boun...@lists.utsouthwestern.edu 
[histonet-boun...@lists.utsouthwestern.edu] on behalf of Vickroy, Jim 
[vickroy@mhsil.com]
Sent: Tuesday, September 02, 2014 10:51 AM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Frozen sections for Immunoflouresence

Lately we are experiencing an issue where the tissue sections are coming off of 
the slide during the staining process.   We have always used positive charged 
slides and have not experienced this problem on a routine case.   We do not fix 
the slides but allow them to air dry for Immunology.  Some have suggested a 
quick fixation in acetone or 95% ETOH.   This doesn't seem to correct the 
problem and we don't find the IF stains work as well.   We have tried a 
different batch of slides and are looking into a different kind of coating for 
the slides.   Our regular frozen sections stay on the slides.  We are using 
OCT.   The only other thing I have noticed is that maybe it's my imagination 
but the OCT in the block surrounding the kidney biopsy seems a little softer 
than normal.   I did try a different mounting medium and maybe the softness was 
my imagination.   Has anyone else experienced this problem and if so how did 
you correct it?   The lab is air conditioned so I don't think the humidity is 
the main factor.  The Immunology staff changed all of their reagents so we have 
reduced that variable hopefully.   We are using Leica positive charged slides.  
(I have found that the Apex control box slides from Leica are a little better 
than the regular Apex slides.   Any ideas?

Jim


This message (including any attachments) contains confidential information 
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are not the intended recipient, you should delete this message. Any disclosure, 
copying, or distribution of this message, or the taking of any action based on 
it, is strictly prohibited.
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Re: [Histonet] Sirius Red (Normal Light) Yellow Colour Degradation

[Grantham, Andrea L - (algranth)]

I've been doing this stain for years and the protocol that I use came from John 
Kiernan (Histonet Archives). If you use Sirius Red F3B (CI 35782) the stain 
will be good for up to 3 years. Don't use a dye that does not have this CI #.
I have used the stain over and over and it always produces excellent results. I 
stain for one hour in the PSR, wash in two quick changes of Acidified Water. I 
usually do not do the hematoxylin step because the researchers requesting the 
stain prefer it that way. The tissue I stain most often with PSR is carotid 
and/or aorta and heart but I have done a beautiful stain on the back of the eye 
including optic nerve. With polarized light it is a thing of beauty to observe.


Andi G

Andrea Grantham, HT (ASCP)
Senior Research Specialist
University of Arizona
Cellular and Molecular Medicine
Histology Service Laboratory
P.O.Box 245044
Tucson, AZ 85724

algra...@email.arizona.edu
Tel: 520.626.4415 Fax: 520.626.2097





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[Histonet] CD34 NHP Tissue

[Joseph Finley]

Does anyone know of a good CD34 antibody for FFPE IHC that works on non-human 
primate tissue - specifically Macaca fascicularis?

Joseph Wayne Finley II
Laboratory Technician III, HTL (ASCP)cm

Department of Comparative Medicine
Medical Center Blvd \ Winston-Salem, NC  27157-1040
(336) 716-1536
jfin...@wakehealth.edu


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RE: [Histonet] Sirius Red (Normal Light) Yellow Colour Degradation

[Bea DeBrosse-Serra]

We always make the solution up fresh. 

Bea

Beatrice DeBrosse-Serra HT(ASCP)QIHC
Isis Pharmaceuticals
Antisense Drug Discovery
2855 Gazelle Ct.
Carlsbad, CA 92010
760-603-2371



-Original Message-
From: histonet-boun...@lists.utsouthwestern.edu 
[mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of André 
Charbonneau
Sent: Friday, August 29, 2014 4:11 PM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Sirius Red (Normal Light) Yellow Colour Degradation

Hello to all, 

This is a quick comment I would like to mention to people completing the Sirius 
Red staining. If you don't seem to have the desired yellow cytoplasm ( I have 
seen in axolotl red blood cells) then the issue might be because you Sirius Red 
liquid is not longer effective. I know this might seem trivial but I was 
reading some previous posts that mention that the solution is good for up to 12 
years. Personally, I have stained several hundred slides with the same solution 
of ~ 250ml of Sirius Red and gradually, the yellow found in the muscle and in 
the cytoplasm in normal light started becoming less vibrant. The solution can 
probably be kept for 12 years but it has limited amounts of usage. Once I 
re-made my solution, the yellow cytoplasm and muscles were vibrantly yellow 
again. This is different from other troubleshooting comments that were posted 
earlier which said to keep the slides in a shorter amount of time in the 
alcohols during the dehydration. Simply changing the Sirius Red would be the 
way to go. Both old Sirius Red and new were compared at the same time using 
same alcohols and acetic acids, thus removing the probability that other 
factors affected the colouring. 

Sirius Red 0.1% in Sat Sol. of Picric Acid: 
Direct Red 80 0.1g
Sat Sol. Picric Acid 100ml


André Charbonneau
M.Sc. Candidate
Faculty of Dentistry 
McGill University




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[Histonet] Frozen sections for Immunoflouresence

[Vickroy, Jim]

Lately we are experiencing an issue where the tissue sections are coming off of 
the slide during the staining process.   We have always used positive charged 
slides and have not experienced this problem on a routine case.   We do not fix 
the slides but allow them to air dry for Immunology.  Some have suggested a 
quick fixation in acetone or 95% ETOH.   This doesn't seem to correct the 
problem and we don't find the IF stains work as well.   We have tried a 
different batch of slides and are looking into a different kind of coating for 
the slides.   Our regular frozen sections stay on the slides.  We are using 
OCT.   The only other thing I have noticed is that maybe it's my imagination 
but the OCT in the block surrounding the kidney biopsy seems a little softer 
than normal.   I did try a different mounting medium and maybe the softness was 
my imagination.   Has anyone else experienced this problem and if so how did 
you correct it?   The lab is air conditioned so I don't think the humidity is 
the main factor.  The Immunology staff changed all of their reagents so we have 
reduced that variable hopefully.   We are using Leica positive charged slides.  
(I have found that the Apex control box slides from Leica are a little better 
than the regular Apex slides.   Any ideas?

Jim


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