AP Call FAQs for Residents

General

Before you ever take call

Please complete the “Resident Orientation Worksheets” for the frozen section laboratory. A lot of what you need to know how to do on call is in those worksheets. Things like how to do certain stains in frozens, how to accession. It’s not an instruction manual, but a checklist for your first week on frozens. It includes a lot of hands on info that isn’t in this document.

Tips for Cutting Frozen Sections

• Don’t over-freeze in liquid nitrogen. Typically 12-13 seconds in liquid nitrogen followed by slow freezing in the cryostat works well.
• Breast sentinel nodes are often fatty and thus challenging to cut. Here are some suggestions from our histotechs:
• Orient the specimen such that lymphoid tissue is near an edge of the chuck, rather than in the center with the fat near an edge. This way the blade cuts into lymphoid tissue first, rather than fat first.
• It’s easier to pick up a section when you have a little extra OCT edge to “pull” on.
• When you’re into the specimen, look and confirm that you’re cutting into lymphoid tissue first. If not, re-orient the chuck so that the blade cuts into lymphoid tissue first.
• Keep the blade cold. Use the gauze to drip liquid nitrogen onto the blade. Per the histotechs, this works better than freezing the chuck/specimen.
• If all else fails, try increasing the thickness of the section. The histotechs don’t usually go above 6-7 microns, but sometimes it takes us residents 8 microns to get something on the slide.

Handling Specimens

Specimen Dropoff

• Surgical specimens: after hours/weekends, have surgeon/clinician drop it off in the front fridge and sign the book.
• Cytology: after hours/on the weekend, transport in regular black top tube (no media) to Core Lab, and they will deliver to Cytology the next working day

RUSH Specimens

• Page/Question: Clinician wants to rush a specimen.
  • Action: Call the histology lab (x4-1425) and provide them with the case number to request a priority change to make sure the case is processed as a Rush and email the RUSHblock email group <RUSHblock@partnershealthcare.onmicrosoft.com>.
  • Note: The case WILL NOT be rushed if the histology lab is not notified. Do NOT change anything in CoPath.
• Page/Question: Clinician wants read of rush case after hours.
  • Action: It is likely the specimen was only recently changed to a rush (if it was originally a rush case, it would have been read earlier in the day). Ask the clinician if the patient is stable, and if clinical management would change overnight based on the results. In most situations, the patient will be stable--tell the clinician that you will notify the attending/resident via email, and they will follow-up the next morning.
• During the week: rushes that are dropped off before ~6-7pm generally are taken care of without a hitch (by accessioners and PAs/small gross techs). If you are paged about a late weekday rush prior to 10pm, you should call small gross (x4-3564) and ask them to gross and process the specimen as a rush.
  • If they aren’t available and/or it is after 10 pm, you should 1) ensure that the specimen is in formalin (unless it’s not a “normal” rush - check the Hub/grossing manuals if it’s a rush you’ve never dealt with before), and 2) send an email to Jenn Patel, Derek Kingman, and/or Jess Houston asking them politely to gross the rush first thing when they get in the next morning.
• On the weekend: Any rush or clinically urgent cases should be accessioned, dictated, and grossed by the on-call resident. You must load all of the rush specimens unless they come in on Sunday after you have loaded the processors.
  • Email the RUSHblock group email to alert histology about the rush case.
  • If you are notified of a rush being delivered after you have the loaded the processors on Saturday or Sunday, contact the clinician. If the clinician is insistent that the results of the slides will change clinical management of the patient and that this cannot wait until Monday, you must gross the specimen, load it into the processor, and notify the on-call attending if a Sunday read is necessary.
  • If it can wait until Monday morning. If the latter, send an email to Jenn Patel, Derek Kingman, and/or Jess Houston asking them politely to gross the Sunday rush first thing when they get in on Monday morning.
  • It is also helpful to send an e-mail to the resident and attending who will be getting the case on Monday to give them a heads-up.

Transplant Cases

• Page/Question: Organ transplant people page you with a heads-up about a prospective transplant
  • Action: Ask when the transplant is happening and ask where the team is now (Ask for the OR number, which will make it clear if they have an OR yet or not, and ask if donor is in OR). Tell them to page you again ~30min prior to the actual procedure. Notify them of our frozen policy between 8pm and 8am, that we will not be in house and need advance notice.
  • Note: The transplant could happen at any time after they first notify you (or not during your shift, or not at all), so to minimize the waste of your time, ask for a more timely page (which may still involve a long wait, but it’s the best option). No matter when they tell you the transplant is happening, it will almost certainly be later than that. That said, if they say the frozen is in 1 hr and they have the OR, it’s best to stick around and be ready.
• When you cut the frozen, cut 2 H&E slides. The organ bank will bring a worksheet for the senior resident to fill out (re: histology of the organ). When they are done, they take one of the frozen slides with them, so in order for us to have an H&E on file, you should cut a second one at the time of the frozen.
• Evaluation and handling of liver allograft biopsies:
  • Freeze the core entirely.
  • Cut TWO slides for each specimen, as one slide will go to UNOS.
  • The most important thing to evaluate is the percentage of macrosteatosis, as this correlates with post-transplant rejection (they like to see <30%).
  • Complete the UNOS form, which includes additional details such as % macrosteatosis, % microsteatosis, extent of lobular/portal inflammation, fibrosis, and necrosis.
  • Retain a copy of the UNOS form for MGH records.
  • Label the UNOS slides with the patient’s name, UNOS number, and specimen type (e.g. right lobe of liver).
  • Leave slides and paperwork at the frozen lab desk for proper accessioning in the morning (do not accession yourself). Leave a note/e-mail accessioning to inform them.
  • Take the tissue down like a normal frozen and put in formalin.
• Evaluation and handling of renal allograft biopsies:
  • Freeze the core entirely.
  • Cut TWO slides for each specimen, as one slide will go to UNOS.
  • The most important thing to evaluate is the percentage of glomerulosclerosis (i.e. the number of sclerosed glomeruli vs. total glomeruli).
    • Note: They need 50 glomeruli evaluated at a minimum, so if you don’t count 50, get a deeper section and add the numbers together.
  • Complete the UNOS form, which includes additional details such as extent of glomerulosclerosis, presence/extent of interstitial fibrosis, tubular atrophy, and arteriosclerosis/hyalinosis.
  • Retain a copy of the UNOS form for MGH records.
  • Label the UNOS slides with the patient’s name, UNOS number, and specimen type (e.g. right lobe of liver).
  • Leave slides and paperwork at the frozen lab desk for proper accessioning in the morning (do not accession yourself). Leave a note/e-mail accessioning to inform them.
  • Take the tissue down like a normal frozen and put in formalin.

Lymphoma Work-up

• In general, if there is sufficient tissue for multiple cassettes, perform a frozen section to triage the material. If you feel that the specimen is too small, page the HP fellow or attending on-call and ask how to handle the specimen.
• See this very helpful information sheet posted above the frozen lab bench, which can also be found on the Hub here.
• Often times, a specimen will come with a requisition that says “r/o lymphoma” but no callback is requested. In these cases, still follow the above guidelines and perform a frozen section to triage the tissue if enough material is available.

Flow Cytometry

• Weekdays: Flow cytometry tissue specimens should be placed in saline and put into the Flow Cytometry box on Blake 3 (near the accessioning area) at room temperature, including overnight. Include completed flow form and copy of requisition form. Log the specimen in the Specimen Send-Out binder.
• Weekends: Flow cytometry specimens should be placed into Streck preservative (vials available in flow cytometry box behind accessioners’ computers) and stored at room temperature (in the flow cytometry box). Include completed flow form and copy of requisition form. Log the specimen in the Specimen Send-Out binder.
  • FYI: Streck claims that samples in their preservative are stable for up to 7 days for flow cytometry analysis, so it should be fine to leave it over holiday weekends.
  • Flow cytometry is NOT performed over the weekend except in emergency cases. Contact the hematopathology attending on-call first to discuss the case. If flow is needed over the weekend, contact Michelle DeLelys <MDELELYS@mgh.harvard.edu>, the flow lab manager.

Products of Conception

• Look up the patient history. Often times, the specimen is for “r/o ectopic”. Notify the senior resident. Float the tissue in a saline-filled Petri dish to identify villi (light microscope in the frozen section lab can be helpful).
  • If villi are definitively grossly identified, notify the clinician and submit one representative section of the villi.
  • If no villi are definitively grossly identified, notify the clinician and submit the entire specimen as a RUSH for permanent evaluation.
• For other types of POCs, contact the clinician to ask what their question is, how urgently the specimen needs to be processed (i.e. over the weekend or Monday morning), and whether additional studies need to be sent (e.g. cytogenetics). You can also loop in the OB attending on-call afterwards via email.

Prepping Specimens

• If the Junior hasn’t prepped a specimen before, he/she should prep them under direct supervision of the Senior. There are detailed descriptions on how to prep all specimens in the grossing manuals on the Hub; however, it is still best for the Senior to support the Junior and be present with them at the bench when they are learning how to prep.
  • Most small specimens: are already in formalin, just put them on the shelf as they are.
  • Most medium sized specimens: gets tossed into formalin except...
  • Fat pad biopsies: for amyloid (half frozen for IF and half into formalin for permanents).
  • Gallbladders: open before tossing into formalin, be sure to put stones in the jar so grossing staff knows they were there.
  • Bowels: Open them before putting them in formalin! Open them before putting them in formalin! Open the ileocecal valve in right hemi-colectomies! Pin out if removed for tumor or suspicious for tumor. Sometimes figuring this out is harder than you think! The pre-existing diagnosis is not infrequently wrong, so use your judgement.
  • Heart explants: weigh, photograph front and back, freeze a piece of left ventricle for IF, and put a tiny piece into EM fixative. Put in formalin.
  • Liver explants: weigh, + photograph, take the margins (hepatic veins and hilar margins = bile duct, hepatic artery, and portal vein), slice at 0.5 cm intervals, stuff with paper towels to ensure fixation. Put in formalin.
  • Lung explants: weigh, photograph and take bronchial and vascular margins. Put in formalin.
  • Breast specimens: Of late the guidelines have been in flux. If at all possible check if a PA (e.g., Donna) is coming in and leave it fresh. If that is not possible consult your senior who will likely run it by the attending on Breast that weekend.
  • Uteri: Weigh, ink (if appropriate), bivalve and + photograph. No need to slice up as you would on frozens, leave it otherwise intact. Put in formalin.
  • Kidney: Check if it’s a medical renal or tumor case. If tumor: weigh, ink, take margins and bivalve, then put in formalin. If medical: ask your senior.
  • Fragmented Leiomyoma: Weigh and add formalin.

Uncommon Biopsies

Fat pad biopsies

• Cut it in half, save half for frozen, and put the other half in formalin.
  • Page/Question: How big should a fat pad biopsy for amyloid be?
    • Answer: Per Dr. Stone, it should be 1.0 cm^3 in size.
      

Cardiac biopsies

• Usually received on ice. Do NOT use a chuck to freeze it. Freeze by putting a drop of OCT on paper or directly into the cryostat and embed the cardiac tissue within it. After the drop freezes, attach the Asset label to the paper (see below), and put it into a small bag with a copy of the requisition form. Scan the Asset label to “SPUF” (SPU Freezer for Frozen Saved Tissue). Place it in the -80C fridge for Warren 5.
  • Apical core biopsy (received fresh, not on ice):
    • Accession the part type as "Heart Exc Left Ventricle" (so it'll be properly processed; using “heart biopsy” generates a different protocol)
    • Save a piece frozen (don't forget the asset label; see below)
    • Save a piece for EM.
  • How to accession an Asset (create an Asset label)
  • FYI: Using a chuck creates a plane between the chuck OCT and the OCT added on top, which creates processing issues.
  • Note: There should be another portion of the specimen for permanents (which is usually accessioned and handled by the accessioners during the day). After hours and over the weekend, if you cannot find another portion for permanents, check with the clinical team before freezing the tissue.
  • After hours or weekend/holiday endomyocardial biopsies for transplant rejection: Per JST, the on call resident should come in after hours to freeze the piece for IF. ### Rush policy regarding endomyocardial biopsies performed the morning of a weekend/holiday - does the normal weekday workflow apply (ie, if received before 2 PM, is processed as a same day rush?)
    

Muscle biopsies

• You should NOT be processing these specimens. Talk to everyone you can (AP Senior, attending, etc.) to ensure the specimen ends up with the right people who are trained to process it. If you are in a situation where you must process it yourself, please refer to instructions on the Hub. [Note: these have been happening more commonly after hours and on weekends and it is no longer uncommon for the resident to process them.]
  

Temporal artery biopsies

• Any temporal artery specimen (regardless of whether or not marked RUSH) should be treated as a RUSH and grossed/processed right away.
  • Do not cut them. Wrap each temporal artery in histowrap, place in a peach cassette with a ribbon, and write “temporal artery” in pencil on the side of the cassette.
  • If a frozen section is requested to confirm if artery is present, shave the two ends of the fragment and freeze the cross-sections. Temporal arteries from Mass Eye and Ear accessioned to Eye Path as Rushes on Fridays, in recent experience, have been treated as Monday rushes, per Eye Path request (ie, they are not re-accessioned to CV for Saturday read). Always a good idea to check in with Eye Path just to confirm though.

Skin biopsies for immunofluorescence

• Biopsies are usually delivered to Pathology in Michel’s transport medium to be processed for immunofluorescence studies. The tissue can be left in Michel’s medium for up to 72 hours (preserves tissue and its antigenicity), so for weekdays and normal weekends, your only job is to make sure the sample is placed in Michel’s medium (do NOT leave it fresh or try to freeze it yourself). The IF lab will process it the next business day.
  • If a clinician calls you looking for Michel’s medium and they are unable to obtain it on the clinical side, we have a small stock either in Blake 354 by the accessioner’s window. The medium can be used even if there is salt precipitation around the lid.
    • Dermatology is responsible for sending their specimens in Michel’s medium--if they ask you for it, let them know that is their responsibility for future reference (but of course provide it to preserve the current specimen).
  • Exception: The ONLY time you should freeze the biopsy yourself is if it’s the Friday afternoon/evening of a holiday/long weekend and it’s possible the tissue will sit in Michel’s medium for longer than 72 hours before it can be frozen by the IF lab. Freeze it in OCT on a chuck (following the same procedure as cardiac biopsies above) and store it in the -80 C freezer for Warren 5.
    • Another situation is If you and the clinician are unable to find Michel’s medium. Don’t panic. Michel’s medium is a transport medium, not a fixative. It preserves tissue for IF for up to 5 days but does not actually do anything to “fix” it. In this case, ask that they personally and expeditiously bring the specimen to you in the frozen lab in saline and freeze it once they deliver it to you.
    • Check with someone more experience if you are unsure about how to orient the specimen for freezing.
      

Skin “jelly roll”

• At night or on the weekend, you may rarely get a skin “jelly roll” with the clinical history of “r/o SJS vs. TEN.” Basically this is a rolled up portion of sloughed skin that should be cut like this in order to evaluate the extent of involvement of the epidermis:
• Rush kidney biopsy for transplant rejection: [draft] During work hours, IR-guided kidney biopsies for transplant rejection are checked by a histo tech for adequate glomeruli at the time of biopsy. Per a discussion with Dr. Colvin, if a weekend biopsy is absolutely needed, the residents can receive and process biopsies (though without assessing for glomeruli first). However, this means that the biopsy may end up being inadequate for evaluation.