Message from the PresidentThe Dog Days of Summer are al-most upon us which means that we have completed another suc-cessful FSH Spring Meeting, held in St. Petersburg, FL. We had a great turnout for the all-day Su-pervisor and Administrator Forum on Friday, and everything else took off from there for the rest of the weekend. Meeting participants attended classes that ranged from General Histology Techniques, Immunohistochemistry, HT Readi-ness, and ended with Making your

Lab Green. Our speakers shared a great wealth of knowledge, and FSH is very grateful for their partic-ipation in the Spring Meeting. The vendor exhibit consisted of a total of 33 vendors showcasing their services. FSH would like to send a huge, “THANK YOU” to the ven-dors who attended. Without their support, our organization would

not be able to provide such a large benefit to our members. As col-leagues and co-professionals of our organization, our membership should take the opportunity to col-laborate with and thank our ven-dors at the FSH Spring Meeting or the NSH Meeting. The upcoming NSH Meeting will be held in Long Beach, California.

“The network connections forged with peers and vendors is truly the key to achieving your professional goals. ”

>> CONT. PAGE TWO

IS32 SUMMER, 2016 EDITION

As the Wheel Turns

Page 1 to 2 President's MessageNSH Announcement

Page 4 to 5Board of DirectorsLetter from Region III

Page 6 to 7FSH Meeting PhotosAwards & Scholarships

Page 8 to 12Microtomy SafetyElectron Microscopy

Page 13 to 14Membership AppCEU Questions

Message from the PresidentI hope those who may not have been able to come to our local meeting will be able to attend this great event, and use it as an opportunity to net-work with your peers, vendors, and elicit new excitement for the many changes that are taking place in our prodigious field. Next year the NSH meeting will be in the Sunshine State in Orlando, FL at the Gaylord Palms, September 15-20, 2017. California may be the West Coast, but Flori-da has the best coast, and we hope to see all of our membership at the 2017 meeting!

It is exciting to reflect on the new knowledge that our membership gained from the Spring Meeting to utilize at their labs. Conversely, the network connections that our mem-bership forged with their peers and vendors is truly the key to achiev-ing your professional goals. This is important if you aspire to grow and move up the ladder in the laboratory or beyond. So, if you have made new contacts, make sure that you foster those relationships and follow-up with them with continued dialogue. Furthermore, nominations are also an important part of our society to

recognize peers and colleagues for their significant contributions to Histo-technology. Since our annual mem-bership renewal will now coincide with our meeting time, we continue to encourage our membership to sub-mit for these awards to nominate, or seek nomination for the awards and scholarships that are given during our annual meeting.

FSH recently had board elections, and the majority of the positions will be continued by the current officers with the exception of the Vice Presi-dent. It is sad to see Michelle Foster step down from her role, but we wish her great success and joy in her fu-ture endeavors. However, I am excit-ed to have Betsy “Dorothy” Woessner as our newly elected Vice President. I have had the honor and privilege to work with this board over the past 2 years, and eagerly look forward to the next 2 years. They all bring great dedication, commitment, and skill-sets to both their work places and to the FSH. I am indebted to them all for their help and dedication to FSH, and would like to express my gratitude in advance for the upcoming term.

I am excited for our profession, eager to see how it continues to evolve, and I hope that everyone will share in my enthusiasm. I am proud of my career choice that I have been part of for 28 years. There is hardly a day that goes by that I am not still learning, and I have confidence in each of you for the same. Enjoy the newsletter!Respectfully,

John ShelleyFSH President2016-2018

>> CONT. FROM PAGE 1

National Society for Histotechnology

Join NSH in Long Beach, California for

their 42nd Annual Symposium. September 16-21, 1016

Visit NSH.org for details.

Pg 2

Poly Scientific R&D Corp.

PresidentJohn ShelleyKissimmee, FLjshelley@fshgroup.org

Vice PresidentBetsy WoessnerOrlando FL bwoessner@fshgroup.org

SecretaryAlice BonaiutoBradenton, FLabonaiuto@fshgroup.org

TreasurerLoretta SaylesIndialantic, FLIsayles@fshgroup.org

Immediate Past PresidentJerry SantiagoOrange Park, FL Jsantiago@fshgroup.org

Awards / Scholarships / Elections Amy ItalianoTampa, FLaitaliano@fshgroup.org

Continued EducationDreama HallLargo, FL 33771dhall@fshgroup.org

Newsletter EditorNisha CrosbyApopka, FL ncrosby@fshgroup.org

HistorianLinda BlackburnSt. Petersburg, FL lblackburn@fshgroup.org

Region I CoordinatorDreama HallLargo, FLdhall@fshgroup.org

Region II CoordinatorBonnie CohenMiami, FLbcohen@fshgroup.org

Membership ChairpersonCharlotte KopczynskiPalm Harbor, FLckopczynski@fshgroup.org

Financial Chairperson

Bylaws Chairperson

Event Planner

Committee ChairsBoard Members

On-Site Quality Microscope Service for all Makes And Models

Clinical, Research, Academic and Industrial Labs

Full Optical Precision Alignment, Gear Lube and Cleaning

Competitive Long-term Contracts Available

Vendormate Friendly

Mobile: 386-627-0012 (Licensed & Insured)

Pg 4

Board of Directors 2016-2018

It is with great excitement that I write my first article as NSH Region III Director! I’d like to thank you for the opportunity to represent our Region. Please know that you can con-tact me at any time for any questions, com-ments or suggestions. Suedotmr@gmail.com

First, I would like to thank those of you that work in our incredible field of Histotechnolo-gy. We must always remember that what we do daily in the lab changes people’s lives!

The NSH mission statement is: “To empow-er the profession of histotechnology through collaboration, education and innovation.”

Currently, there are 391 members in Region III. We as members need to continue to spread the word to others about the opportunities and advantages that membership has both at state and na-tional levels. As histologists we need our voices to be heard and carried.

Have you signed in on the NSH site, “The Block” yet? If not, you’re missing out on the Society’s pri-vate community. This is where members can connect with colleagues, share knowledge and access resources. Check it out, it’s just one of the perks for being an NSH member.

This year our region has been very busy with state meetings. I have attended two meeting so far this year. • I attended the NCSH spring meeting, held April 7th-9th. The meeting was well attended and had a nice variety of workshops. • The Georgia Histopalooza meeting was April 22nd-24. What a beautiful location and a great opportunity for some good workshops. • Florida had their meeting in St. Petersburg, May 19th-22nd. Unfortunately I was unable to attend but from all the Facebook postings its look like everyone had a great time. • Upcoming is the Mississippi meeting July 16th. This is a one-day meeting in Flowood, MS • South Carolina has their state meeting planned for November 4-6th at Litchfield Beach and Golf Resort, Pawley’s Island.

I would like to send a special thank you to those of you who have coordinated and planned these meetings. I would also like to thank the vendors and sponsors who help support these meetings. Without you it would be difficult or impossible to provide these meetings for our members.

In closing, I want to remind everyone about the NSH Symposium/Convention coming up. This year’s 42nd annual S/C will be held in Long Beach, CA, September 16-21, 2016. I hope to see you there.

Respectfully,Sue J. ClarkNSH Region III Director

Pg 5

Annual MeetingPg 6

Thanks to all of our members and vendors who attended our 2016 Annual Meeting and made it a huge success! We look forward to seeing you all again in 2017.

AwardsEvery year, FSH awards special recognition to our members and peers. The Board of Directors, after careful evaluation of the submitted nominations for the awards, decides on the recipients of the awards given at the spring meeting.

Here are our award winners:

Histotechnologist of the YearJodi Balsi

Career Achievement Award

Isis Gentles

"Bobbie Spillan" AwardJerry Santiago

Vendor Appreciation AwardJoe Myers from BioCare

President's AwardLoretta Sayles

Pathologist of the Year AwardDr. Charles Perniciaro

ScholarshipsThe Florida Society of Histotechnology is awarding (3) $500 scholarships sponsored by Poly Sci-entific and FSH. This year Allied Search Partners has added a fourth scholarship in the amount of $250. These scholarships are awarded on a reimbursement basis.

Poly Scientific R&D Educational Scholarship Aidyn Quintana Alvarez

Excellence in Histology Educational Scholarship, sponsored by Poly Scientific

Chelsea Nagle

The FSH Education Scholarship

Jaritza Sweat

Education Scholarship from Allied Search PartnersSamaan Samehrship

not pictured

Pg 7

At this year’s FSH meeting in St. Petersburg I had the pleasure of giving a presentation entitled, "What Every Microtomist Should Know!” The presentation consisted of discussions of microtomy basics, practical microtomy applications, safety, and ergonomics.

The information I found when researching safety was neither up to date nor precise or practical. I wanted the attendees to be able to take home a list of Microtomy Safety Precautions. To this end I made our safety discussion a group activity.

The attendees where broken up into two groups and instructed to prepare a list of 10 Microtomy Safety Precautions. The following list is a result of their work.

1. Make sure all screws and clamping devises are secure. 2. Handle Microtome blades carefully when putting blade in or taking out from blade holder. 3. Always use knife guards when appropriate 4. Store blades new and used in covered container 5. Brush AWAY from the blades edge when cleaning 6. NEVER place your hands or fingers above the blade 7. Report malfunctioning microtomes 8. Never leave blade in knife holder when microtome not in use 9. When leaving microtome for the day remove blade and lock fly wheel 10. Take breaks!

Thanks to those who participated in preparing our list of Safety Precau-tions:

Patricia Lewis, Olivia Macias, Leesha Moore, Michelle Newman, Lourdes Rivera, Elba Vidal, Lindsay Vail, Dor-othy Woessner, Alice Bonaiuto, Iliana Laserna.

Microtomy Safety Precautionsby Bonnie Cohen, BS. HT(ASCP)HTL

Pg 8

usdiagnostics.roche.comwww.ventana.com

VENTANA is a trademark of Roche.© 2016 Roche PP-US-06659-0316

We don’t just see slides with tissue samples. We see real lives touched by cancer, and incredible stories of hope.

For this reason, our single focus is to advance patient care. We’re persistently innovating and finding new ways to integrate technologies. So today, we’re proud to offer the power of the VENTANA portfolio as part of our end-to-end offering as Roche Diagnostics.

Together, let’s create stories of success for more patients than ever before.

EVERY SLIDE TELLS A STORY

That’s why we’re bringing advanced solutions to life

TD_Brand_AD_Every Slide_ARCHIVES.indd 1 3/23/2016 4:57:03 PM

Transmission Electron Microscopy (TEM)by George Price

Many histotechnology training programs have a requirement to gain an understanding of electron microscopy (EM) as part of the curriculum.

Some programs have students visit an EM lab to gain an understanding of the techniques in-volved in producing samples suitable for diag-nostic evaluation with the electron microscope. Where others may require an understanding of how the techniques are achieved through the re-view of written material.

For students visiting my lab, I try to demonstrate how tissues are processed, cut, and stained for TEM in relation to the techniques used to pre-pare histological samples for light microscopy. The rationale is the same, but different materials are used for TEM preparation.

Fixation or preservation of tissue samples is the first step. For EM, glutaraldehyde buffered with sodium cacodylate is preferred as the primary fix-ative. Glutaraldehyde crosslinks proteins better than formalin, and results in better preservation at an ultrastructural level. Formalin fixed tissue can be used for TEM, but the combination of less preservation and phosphate buffer of commer-cially available formalin results in less contrast of the TEM image.

Tissue sample size for TEM should be no larger than three cubic millimeters, allowing for good primary fixation, and for good secondary fixa-tion with buffered osmium tetroxide. Osmium fixes lipids, providing good fixation of membrane structure, but will only penetrate tissue one and a half millimeters. With the osmium fixation pen-etrating the tissue from all sides, tissues greater than three cubic millimeters will be left with un-fixed tissue in the center.

Osmium is a heavy metal found in the platinum group on the periodic table of elements (atomic number 76). Besides its ability to fix lipids, os-

mium will start to impart contrast to the tissue viewed in TEM. Since TEM uses an electron beam instead of light, other heavy metals of ura-nium and lead salts are used to impart the con-trast one sees in the final stained thin section to be viewed.

All of the elements that make up biological samples (carbon, hydrogen, nitrogen, oxygen, phosphorus, and sulfur) are not heavy enough to scatter an electron beam, hence the need to impart contrast to the tissue sample with heavy metals.

The next step will further impart contrast to the tissue sample through the process of en bloc staining. This is achieved by immersing the sam-ple in a uranyl acetate solution; uranyl acetate is also the first of two heavy metal stains used on finished thin sections to be viewed on the TEM.

This is followed by dehydration with ethyl alco-hol, and then replacement of the alcohol with an intermediary solvent before infiltration with epoxy resin. These steps are similar to routine histological processing:

1. Alcohol removes water from tissue since paraffin is not miscible with water.2. Alcohol is replaced with the intermediary solvent xylene since paraffin is not misci ble with alcohol.3. Molten paraffin replaces the xylene in the infiltration step.

Ethyl alcohol is used as a dehydrating agent in EM tissue processing because it causes the least amount of clumping of proteins in the cell cytoplasm than other alcohols, resulting in better ultrastructural detail.

The intermediary solvent used in EM processing is propylene oxide since it dissolves epoxy resin quickly and completely.

Pg 10

>> cont.

It is also extremely volatile, and will evaporate over a number of hours leaving the epoxy resin completely infiltrated into the tissue.

After infiltration, the tissue is embedded with fresh epoxy resin in disposable polypropylene capsules that are made for this specific purpose. Although the epoxy resin has a catalyst, or ac-celerator to facilitate its polymerization, heat is applied to speed up the polymerization process.

Once the resin has polymerized, or hardened, the polypropylene capsule can be cut away leav-ing the finished tissue block ready for microtomy.

Using a razor blade, the tissue block is trimmed to expose the embedded tissue, and then placed in the tissue chuck of the ultramicrotome.

Glass or diamond knives are used to produce sections of tissue on the ultramicrotome. Glass

knives dull, or pick up knife marks quickly, and are labor intensive to make and use. Glass knives are relatively cheap, but for quick and re-liable results diamond knives are best.The knife is placed in the knife holder of the mi-crotome stage. There are adjustments to the knife holder and chuck to align the block so it is perpendicular to the knife. The block of tissue is “faced” in much the same way paraffin blocks are, but the sample size is much smaller and the alignments and cutting are done under 10x mag-nification of a stereo microscope.

Once the block is faced, a trough that holds wa-ter behind the knife edge is filled; this trough is referred to as a “boat”. The epoxy is somewhat hydrophobic, allowing the subsequent sections to float off onto the water surface of the boat. Thick sections ½ to 1u are cut and transferred to a drop of water on a glass slide and dried on a hot plate. The thick sections are then stained

with the monochro-matic stain toluidine blue-o, and then ob-served with a light microscope to estab-lish adequacy of the sample. In the case of kidney samples, glom-eruli are required to be in the sample for diag-nosis.

Once adequacy of the tissue sample is estab-lished, thin sections are cut. These thin sections may range in thickness from 20 to 80nm in thickness. The section thickness is determined by the

Transmission Electron Microscopy (TEM)>> cont. from page 9

Pg 11

>> cont.

Transmission Electron Microscopy (TEM)>> cont. from page 10

Pg 12

the interference color of the section floating on the water surface; 20nm being a silver color to 80nm a dark gold.

The thin sections are then transferred to small three millimeter diameter screens called grids. The thin sections are then stained first with ura-nyl acetate and then lead citrate. This so called “double stain” is what adds the final contrast to the tissue section to be viewed by TEM.

As delicate as the work is to produce samples for TEM, there is one advantage to staining for EM over staining for light microscopy: The embed-ding media does not have to be removed from

the tissue for staining. The stain is simply placed on the grid for two minutes for each stain and then washed off.

The stained grid is then placed in the TEM for ob-servation and documentation with photographic or digital imaging.

by George Price

CEU QuestionsTransmission Electron Microscopy

Instructions: Circle the BEST answer.

1. Fixation is the first step in preparing tissue for electronic microscopy. This is accomplished by a two-step process. What two fixatives are used? a. Formalin; Osmium tetroxide c. Glutaraldehyde; Formaldehyde b. Zinc; Osmium tetroxide d. Glutaraldehyde; Osmium tetroxide

2. Fixation with _____________ results in less preservation and contrast of the TEM image: a. Phosphate buffer c. Glutaraldehyde b. Formalin d. Alcohol

3. Osmium fixes _____________ by providing good fixation of membrane structure, but will only penetrate tissue one and a half millimeters: a. Muscle c. Lipids b. Nerve d. Keratin

4. En bloc staining is achieved by immersing the sample in a _____________ solution: a. Phosphate buffer c. Acid alcohol b. Osmium tetroxide d. Uranyl acetate

5. What types of knives are used in EM processing? a. Diamond d. A and B b. Glass e. B and C c. Steel

6. Epoxy is considered hydrophilic, allowing the subsequent sections to float off onto the water surface of the boat. a. True b. False

7. Ethyl alcohol is used as a dehydrating agent in EM tissue processing because it causes the least amount of clumping of proteins in the cell cytoplasm than other alcohols, resulting in better ultrastructural detail. a. True b. False

8. Epoxy resin has a catalyst, or accelerator to facilitate its polymerization, light is applied to speed up the polymer ization process. a. True b. False

9. What infiltration media is used in EM processing? a. Paraffin c. Acrylic b. Epoxy resin d. Paraplast

10. What two reagents are used to “double stain” sections for the final contrast for TEM? a. Propylene Oxide d. A and B b. Uranyl Acetate e. B and C c. Lead Citrate

Dreama Hall, FSH CEU Chairperson Cost: FSH Member – FREE; Non-Member - $10.00 1155 Lake Avenue, S.E. Mail your answers and applicable fees to: Largo, FL 33771

Pg 14

Nominate a deserving co-worker for an award.

Nominate a deserving student for a $500 scholarship to help them succeed.

Did you know?You can renew your membership online! www.fshgroup.org

We want to hear from you. What’s new in your lab? Do you have an opinion? Send us an article you feel would be of benefit to your fellow Histotechnologists. Do you have a question, a tip or a comment? It’s easy, just contact the FSH newspaper editor for consideration.

It’s time to renew your FSH membership! We want you to be a mem-ber of the only professional organization for histology professionals in Florida. As a member, you will receive:

• E-Newsletter posted on the FSH Website• Continuing education opportunities at regional and state meetings at a dis-

counted rate. • Self study continuing education opportunities through our website offerings• Access to updated profession information on the FSH website at www.fsh-

group.org, including links to NSH, ASCP, and Clinical Lab Board• Continuing Education Credits provided by FSH free of charge• Representation at the Board of Clinical Laboratory meetings• Representation in the House of Delegates during the National Society for

Histotechnology Symposium/Convention• Updates from the NSH Regional Director

The annual membership dues are $35.00 and membership runs from April 1 to March 31. To renew, please visit www.FSHgroup.org.

Est. 1973

SUMMER, 2016 EDITION

As the WheelTurns