Periodic problems with FNAB smearing

RE: Periodic problems with FNAB smearing

William Easson — Sun, 15 Jun 2025 19:34:39 GMT

The 'more and more' phrase is because I left vet school in the 90s being a dab hand at FNABs, and reports would periodically comment on the good quality. More recently (in the last year or so) I had noticed that as I drew the (perpendicular) spreader slide along the sample slide, it got pushed/stuck to the sample slide. I wasn't exerting the pressure, I think the spreading was evacuating the air gap between them and the atmosphere was pushing them together.

I just had an MCT reported with good preservation, so the slide rubbing is working for n=1.

As I write this I think this may coincide with a practice I'm working at - perhaps their brand of slide is different (they are frosted, non-bevelled, but do feel somewhat scratchy when they touch one another).

Smears I've found easier by rubbing a paper towel on both slides, and varying the angle between them depending on the size of the drop to include more or less blood in the head.

RE: Periodic problems with FNAB smearing

William Easson — Sun, 15 Jun 2025 19:28:04 GMT

Thankyou very much for all your advice!

I've found some change by rubbing both of the slides with a paper towel beforehand. I had been taking them straight out of the packet. I've also found, weirdly, that doing this has restored the curved feathered edge to my blood smears. Whether this is cleaning them, or putting a coating of something onto them I don't know. Time and FNAB reports will tell if the cells are better preserved, but for now they aren't being pushed together by negative/atmospheric pressure any more.

RE: Periodic problems with FNAB smearing

Roger Powell — Tue, 10 Jun 2025 12:41:14 GMT

Yep, use that too and it's great!
Removes greasy finger prints too (which can interfere with even dispersal, contaminate etc) as well as useful for cleaning your scope and oil transfer to your x40! Just a pain to keep doing it for this and you have to wait for it to dry (appreciably seconds) and when it shouldn't really be required if they are stored well and maintained.
Would use it though if you happen to sneeze when sampling etc as that can mess up parts of the smeared material too!

Spreader slide for blood smearing should be treated differently but cleaned with dH20 in between smearing (cheaper and safer, as good) but again replaced and remade as required when checking smear quality.
For cyto, the spreading should transfer all the cells across/onto the upper slide and you can check cellularity and preservation directly when wet/still drying and critically unstained, via the scope and a lowered condenser, very easily without any staining required (and waiting for it to dry beforehand), to then repeat the sampling and smearing while the animal is still there/under anaesthesia etc.

Volume wise, I would stick to about 5mm max. diameter cellular droplet/puddle to allow for sufficient spreading, as otherwise you are best separating the droplet/aspirated material and spreading smaller volumes well on multiple slides, rather than 1 larger droplet that is still too thick or goes off the ends/sides.

Not yet worked at a lab where our threshold for slide rejection has lowered, but an interesting thought and proposal - still trying to get consistent Hx and actually smeared preps, rather than just squirted droplets!! (-:

RE: Periodic problems with FNAB smearing

David Scarff — Tue, 10 Jun 2025 08:57:28 GMT

Cleaning (I use isopropyl alcohol) and drying the slides also helps as if they are damp at all this increases the surface tension between the slides which can crush the cells. Also if possible stain the residue on the spreader slide so can check how the cells look. Make sure there is only a small amount of material on the slide as this produces a better smear.

I also think the threshold for slide rejection has possibly lowered at the labs.

RE: Periodic problems with FNAB smearing

Roger Powell — Mon, 09 Jun 2025 16:43:52 GMT

Hi William,

Quite a few factors in this and not sure all of them are in your control(!), as your outlined approach sounds fine and I don't really have enough of the info on "more and more" etc, or any unnoticed change in your smearing....?
Some thoughts to hopefully help are:

With the "drawing", are you going 'parallel' or 'perpendicular' as the latter is easier to control, so the only forces spreading the cells come from capillary action and your pull, no downward pressure applied, with all the cells typically then transferred to the upper slide.
If you're finding cells are remaining on the lower slide and especially nearer its edge, that's typically because the drawing is (becoming) angled slightly, so the scraping over the edge damages/lyses the cells, for "poor preservation" then, even if diagnostically cellular.
If the slides actually stick when pulling apart, it can result from too much liquid and inevitably, as you more forcefully separate or continue the draw, then ruptures and lyses the cells.
How 'attracted' the slides are to each other can also be a factor that is hard to overcome / compensate for here, as it results from the quality of the slides and glass (presuming uncoated), seemingly worse when using non-frosted ones, non-bevelled, cheaper glass....
You can get a feel for this being likely to happen (and possibly then a/the factor in your situation?) if the slides/practices you're getting "poor preservation" correlate with slide manufacturer or batches where they were hard to separate when getting them out of the box?
Similarly, if they get a little damp, this will increase the chance of them sticking together (in the box and when used) and directly lyse the cells too.

Particulate matter like Rob says will cause uneven pressures and focal lysis when drawing but shouldn't result in all the cells being mushed, as with invisible glass shards (again from poor quality slides).
Wiping them down pre-emptively helps, as outlined, if they're not being stored properly/getting dusty/damp etc, and when you do this 'cleaning wipe', again if you find the lint fabric/tissue paper is sticking and hard to wipe along/across, can also indicate the slides won't then spread well and will damage the cells as they stick together etc
The use of additional suction (cf. just capillary action into a 'jiggled and spun needle' for most skin masses etc) as Rob says too is a factor, more often haemodiluting though, unless also compounded by using a smaller/too small gauge needle (eg 25G below) but again would seem unlikely to have changed for you?

Cannot emphasise how important it is to spread the cells out, so thank you for that (from a cyto perspective!!), and while I would always recommended what you are/have been doing, maybe alternatively try out a similar haematology 'pull-drag' technique then now, but with a shallower angle and slightly slower (cf. blood smears) - again though, no down ward pressure, just resting the angled and fore-shortened spreader slide on the top to pull back to the droplet and then drag it forward.....
Some people find this works well for them, but personally find/found it does rupture the cells more often as the forces at the leading edge are greater, so delicate cells (eg. lymphs) will more often get damaged, in part as there is less liquid too in aspirates comparatively to blood.

A final thought (which may be artificially accentuating/biasing what appears to be going on) is that you are (also) seeing the vagaries of a cyto report as we do not (unfortunately - sorry!) have a consistent and standardised way of grading the preparations, that sentence varying from pathologist to pathologist, even within a pathologist as they unknowingly apply different criteria to different sample types......
Can't comment on that further, but would add here that if your technique can smear a lipoma harvest well and intact, it is fine and very good, so stick with it (and personally would say thank you, as those are (a nerdy) work of art!! (-:)

Good luck but let me know if I can answer anything more specifically, or not making sense with the above!

Best wishes,
Roger.

RE: Periodic problems with FNAB smearing

Rob Loxley — Sat, 07 Jun 2025 15:24:07 GMT

Give the slides a quick wipe clean 1st - sometimes bits of dust seem to rupture cells. Also excessive (sometimes any) aspiration during collection will rupture cells before even spread.