Knowledge Lab

Welcome to Tails from the Lab, where an ever optimistic veterinarian and slightly salty technician entertain listeners with true stories and tall tales revolving around laboratory diagnostics.

Names have been changed to protect the innocent, but the lab work is real.

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Each episode we hope to leave you a little smarter, a little brighter, and feeling more empowered in the lab.

Hello. Hello. This is Jessica and this podcast is Tales from the Lab. Today we have a special episode which is one of our tech talks. These are short episodes that will contain information that will allow you to perform some of the diagnostic tests that we have mentioned in our larger full episode podcast.

Today’s tech talk topic is the fluid analysis.

So we’re going to cover how do we get the sample,

what tubes do we put it in, what can we do with those tubes? And then just, you know, a couple of differences between the in house analysis and the reference lab.

So our fluid samples are obtained primarily through abdominal and thoracosentesis. So we are doing this ultrasound guided. So the clinician on the case will identify a pocket of fluid and then obtain that using an ultrasound.

And this is really nice because we can actually visualize really small pockets of fluid because even a drop can give you a lot of information.

I do want to kind of point out one little tip in here.

And we learned this really when Dr. Brown came to the hospital and we started to do our own in clinic fluid analysis. She would notice that there would be this kind of magenta material on our cytology slides and also in our fluid samples.

And what we realized was happening was the doctors were using ultrasound gel as part of their ultrasound. And it actually causes an artifact once it is stained. And it doesn’t kind of matter how many times you wipe the patient off.

Any type of ultrasound gel that’s used is going to cause an artifact on your slides, which can make it hard to identify things like bacteria or certain types of tumors.

So we want to try to stay away from using ultrasound gel when we’re knowing that we’re going to get, you know, a cytology sample, whether that’s cytology from a mass or whether we’re going to get a fluid sample.

So just a kind of little tip and we’ll go ahead and put an image of that up on our Instagram. Tails from the lab. That’s tails spelled like dogtail.

Another option we have for obtaining fluid samples, especially in patients that have had some type of surgery, is using if you guys have a Jackson Pratt drain in place, you can use that fluid that is being collected as a fluid flu for your fluid analysis.

And this is really nice for trending options. And we’ve talked a lot about trending throughout many of our cases that it’s important to be able to see where things are going.

And we use this like if we have a foreign body surgery that has a Jackson Pratt drain in, we’ll go ahead and watch that fluid kind of change over the course of several days and we can use that as our way to gauge whether, you know, our antibiotics are working,

make sure that we don’t need to go back in for surgery,

and really allow the doctors to have a better idea when to go ahead and pull that drain.

So we’ve identified our fluid sample either on ultrasound or if we have a Jackson Pratt drain in place. Now, what do we do with that and what tubes do we put that in?

So there are going to be two types of tubes that we collect fluid in. The first one, and probably the more important one is going to be an EDTA tube.

The second one, which I like to think of as a bonus tube, is going to be a non additive tube. So it’s going to be a red or a white top, depending on the manufacturer of your tubes.

So we’ll talk a little bit about the EDTA first tube first and then we’ll come back to the non additive tube. So your EDTA tube is going to do two things.

One, if you have a particularly hemorrhagic fluid, it’s going to prevent clotting of the sample and, and then it’s also going to help preserve cell morphology. So the EDTA that’s in there is going to help those cells stay nice and healthy.

This is especially important if you’re sending that sample off to the reference lab because it’s going to not be fresh. So our non additive tube, the red or the white top, is going to be used for things like culture biochemistry testing.

So this is, like I said, a bonus tube. If you have enough sample, you can go ahead and fill one of these up so that you have it just in case.

And as far as sample volume, we want to go to the fill volume. So if you have an EDTA tube that requires two mls, you want to just fill it up with two mls.

We don’t want to overfill our tubes, just like we don’t want to overfill them for blood samples.

So a fluid analysis contains a couple things. It contains visually looking at our sample for Color and tubidity, protein concentration, a nucleated cell count, plus or minus a red blood cell count or a PAC cell volume and then also the microscopic analysis.

And we’re really just going to focus on those first three and then we’ll go in more depth for the microscopic analysis at a different time. So protein measurement, how do we measure protein in fluid?

So we can do it a couple different ways. We can use our refractometers, which is what we use for our urine specific gravity, or we can use our chemistry analyzer.

So for refractometry. On our refractometer we’re just going to place a small drop of fluid just like we would for urine specific gravity, and then we’re going to take a peek inside there and oftentimes there’s going to be your urine specific gravity measurements and then there will also be a serum or plasma protein measurement and that’s what we’re going to obtain.

So we’re going to look at, at that series of measurements, not the urine specific gravity. If we are using our chemistry analyzer, we’re just going to take that total protein slide that comes with, you know, our chemistry, our in house chemistry analyzer and we’re going to measure the protein off of that.

A little tip about protein measurements that can help. A lot of times our fluid samples have some type of color to them. So maybe they’re ******, maybe they’re a little bit milky, maybe they’re pink.

If you take and spin down your sample in a stat spin or a centrifuge, just like you would for a urine sediment slide, you can take the supernatant that’s on top of the kind of pellet that’s been condensed at the bottom and you can use that supernatant, that liquid on top,

as what you are using to measure your protein. And getting that kind of color out of there can oftentimes make that line crisper on the refractometer to measure the protein in there.

And it can also help your chemistry analyzer, which oftentimes is based on color change,

to not have a lot of color in the sample. So that’s just a little tip that can help with measuring proteins. Okay, so we talked about our protein concentration. Color and turbidity is important, I think, especially if you’re going to be sending maybe a cytology slide to the reference lab to include.

So we’re going to describe the color of the sample and then we’re also going to describe, you know, is it clear, is it cloudy, are there Big chunks floating inside of it.

So that covers the color and turbidity portion.

A nucleated cell count. So how do we do that? The easiest way to do that is to actually use our hematology analyzers, measure fluids in there and take what would be your D WBC count, so your white blood cell total count, cell count, and use that as your nucleated cell count.

And Dr. Brown likes to say hematology analyzers are cell counters. That’s what their job is.

So they’re really good at counting those cells. So we’re just going to take that total white blood cell count and that will be our total nucleated cell count. And why are those things important?

And if we think way back to tech school or to, you know, vet school,

our, our professors probably talk some about fluid classifications.

So things like transudate, modify, transudite and exudate. So these are kind of the buckets that we start to put our fluid samples in. So they will help the doctors with kind of their differential diagnosis.

So the things that they need to kind of put them into those buckets of the different categories of fluids are going to be your total protein measurement and a nucleated cell count.

So that’s why having those measurements is super important.

So what do you do if your supervisor doesn’t want you to put your fluids into your hematology analyzer to get that white blood cell count for your total nucleated cell count.

So then we have to move on to slide preparation.

So there are two types of slides that we like to prepare consistently for our in house fluid analysis. And that’s going to be called what’s a direct slide and then also a sediment slide.

And so our direct smear is just like a blood, a blood film. So we’re just going to take a drop of our fluid out of that purple top. We’re going to place it kind of at the back of the slide.

And then just like we would we would for a blood film, we’re going to take our spreader slide, pull it back into that drop, having it somewhere between a 35 to 45 degree angle.

And then we’re going to push out so we get that kind of nice feathered edge. And this direct slide is going to be the one that you can use for a manual total nucleated cell count.

The sediment slide is a little extra slide that we like to use, especially when we have a cell count that’s super low.

One of those modified transudates or even a transudate effusion we want. We will. We can spend a lot of time looking around on the direct smear for the cells that are there.

So the sediment slide just goes ahead and concentrates those cells so that we can see them a little easier. We don’t have to do as much search, searching. So again, you’re going to use that stat spin and your little Eppendorf tube.

You’re going to put a small amount of your ed a fluid sample into that stat spin tube and you’re going to spin it. We usually spin it on our normal setting and then pour off the supernatant.

Or if you want to use it for your protein measurement, you can use that.

And then we reconstitute that pellet of cells that’s at the bottom of this of the tube and take a transfer pipette. And just like we would for the blood film, we take a small drop, we put it towards the kind of the back of the slide.

And then we’re going to take our spreader slide again and we’re going to pull back into our sample that we have there. We’re going to push out. We’re just going to go about three quarters of the way out and that with our spreader slide and then just pick straight up.

And what this produces is a line prep. And so it’s called a line prep. And again, there’ll be an image of this on Instagram, the difference between a direct and a sediment slide.

But that line prep slide has a line of cells that are concentrated across the top. And that makes it really easy, a nice, easy place to look for those cells that maybe we would have had to search all over the body of our direct slide for.

So it’s just a nice quick way to be able to concentrate those cells.

That’s really the end of our fluid analysis. Our in house fluid analysis. We talked about color and turbidity. So describing the fluid, because maybe the clinical pathologist at the reference lab isn’t going to get to be able to see that.

We talked about the protein measurement. We can either use our refractometer or we can use our chemistry analyzer. And then we talked about that nucleated cell count. So the easy way is to put your fluid into your hematology analyzer.

And then in another episode, another TechTalk episode, we’ll talk about how to do an estimated nucleated cell count using that direct that we made.

We also talked about the two types of slides to make the direct and the sediment and what the differences are for both of those So I hope you enjoyed this brief tech talk and we will talk to you guys soon.

Have a great day.

Tails from the Lab is a production of Antec Diagnostics. The intent of this podcast is to provide education and guidance with the understanding that any diagnostic testing and treatment decisions are ultimately at the discretion of the attending veterinarian within the established veterinarian patient client relationship.