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A Mayo Clinic podcast for laboratory professionals, physicians, and students, hosted by Justin Kreuter, M.D., assistant professor of laboratory medicine and pathology at Mayo Clinic, featuring educational topics and insightful takeaways to apply in your practice.
- This is Lab Medicine
Rounds, a curated podcast
I'm your host, Justin Kreuter,
a transfusion medicine pathologist
and assistant professor
of Laboratory Medicine
and pathology at Mayo Clinic.
Today we're routing with Dr.
Brooke Katzman, assistant professor
of laboratory medicine pathology
and consultant in the Clinical
Core Laboratory service
division at Mayo Clinic.
Thanks for joining us today, Dr. Katzman.
- Thanks for having me.
- You know, so we wanted
to have you here to talk
about kind of the rationale
behind blood tube collection order.
And I know before we started
recording, I was confessing
to you that, that I
oftentimes am struggling with
what color top do I need for what tests.
And so I'm really jazzed to kind
of get this a little bit
straightened out with you maybe
for our audience, since
we're really a diverse
audience here.
Why is it important that kind
of different sample collection
tubes are collected in
a particular order?
- Sure. Yeah.
So the, the majority of our
collection tubes have some sort
of additive added to that particular tube.
And so really that's what it
comes down to is being able
to make, make sure that those
don't cross into other tubes.
So for example, when a
needle's inserted into the tube
there, there's a slight chance that
that ne needle will pick
up some of that additive.
So for example, blood
splashing against the needle.
And then the issue is then when
that next tube is collected,
there's a risk that it'll carry
over into that tube leading
to a contamination by that additive.
And this, I would say, is more
of an indirect contamination
where it's, it's happening
again because of a, a splash
or, you know, transfer from the needle.
One example of maybe more
of a direct contamination
would be if a pH phlebotomist
realizes that, you know,
they collected the tube out
of the wrong order, let's
say for example, EDTA.
So potassium EDTA is one
of the additives in
our light purple tubes.
And so rather than restarting,
re poking that patient,
they then pour
that tube over into the
SST tube, for example.
And so now you have a serum a
tube that now contains EDTA.
And so you go to test that and,
and now you have some
really discrepant results.
So both of these scenarios,
test results for some
of those analytes, they're
gonna either be falsely elevated
or falsely decreased.
And, you know, we know that has a,
has a direct impact on patient care.
So you risk misdiagnosis mistreatment
often re results in a
recollection of a specimen due
to a spurious result, whether
it's detected by the lab
or detected by providers
where it's just not consistent
with clinical picture.
So really to avoid this,
the Clinical Laboratory Standard
Institute, which is CLSI,
they've set forth a a
standard recommended order
of collection within their guidelines.
And this is really what all laboratories
and phlebotomy units are
supposed to be following.
- You know, I I I'm glad you
kind of brought up this idea
of kind of this, I guess you
were talking about as direct
or indirect contamination, right?
So, you know, I'm sure there's some
folks in the audience where
yeah, they could understand, oh,
I collected the wrong tube and,
and sort of innocently
feel like, oh, let me put
that into the right tube.
Not appreciating that there is,
there are particular additives
in these different tubes
and how that could actually
cause an interference in,
in the lab testing, like
you're saying, which, you know,
there's gonna be additional costs, right?
If the test needs to get
right, rerun, potentially
inappropriate care for the patient,
if somebody's acting on false results
as it may not get picked up.
If it is picked up, there's
even a delay then to get
that additional testing.
So I guess what you're saying is like,
when we make a mistake, we just need
to call foul it ourselves and Yeah,
- And you know what, often it's in the
best intentions, right?
A phlebotomist doesn't
wanna restick a patient,
they wanna say, you know,
I already have this blood,
why can't I use this, put it in this tube.
So that's, that's the challenge.
It comes from a good place,
but we just have to, I
think it's important just
to realize why exactly we're
doing what we're doing.
- Yeah. And to understand how, you know,
gratefully it's nice that when
we have phlebotomist, people
that are trained in to do
this, to take this role,
but I know nurses also
train in and do phlebotomy.
I, I remember when I was in
medical school, I, I did a,
a number of collections
from my own patients.
So, so we have a lot of people on the team
that may be involved and,
and this is important
knowledge to understand.
So I think we're gonna dive in.
What, what is the particular order?
Can you kind of like walk us
through that, that rationale?
Because I think that might
get at that point about
that concern about potential carryover.
Like why is the order of the rainbow
- The order the way it,
that it's, yeah, yeah, yeah.
So this is kind of a,
I'll just say is a very
simplified version of really
what the intent is is or what,
what the differences are.
So the first collection would
be a, a blood culture tube
or a blood culture bottle.
And, and those are, you
know, those are gonna be for
specimens to, to measure
bacteria, identify bacteria.
We want basically the cleanest specimen
that we can for that.
The next tube is what we
call our light blue top
or our sodium citrate tube.
These are tubes that are often
used for coagulation testing.
And just another other important
pre-analytic component is
that they have to be completely
filled to ensure that
that proper ratio of blood
to anticoagulant sodium
citrate in this case
is the anticoagulant.
The next group would be the serum tubes.
And those could be those with
or without a clot activator or a gel.
And so we think of, excuse me,
sssts serum separator tubes.
They may have a gel in
it that once you spin it,
it separates the blood from the serum.
They can be gold in color,
but these tubes are typically used
for general chemistry testing.
The next are the heparin tubes.
So these tubes can again, also be
with or without gel.
They can be either dark
green, light green tops.
Typically, again, chemistry
tests are performed
using these specimens.
This is gonna result in plasma.
And we specifically use
these in STAT testing so
that we can rapidly test the samples.
The, the difference with the
serum is that we need it to,
we need about 30 minutes
for that specimen to clot
before we can test that sample.
So with plasma we can test immediately.
The next tube type would be
the EDTA potassium EDTA tube.
This would be with or
without a gel separator.
Again, these are often lavender
or light pink, purple in color.
And these are gonna be used
primarily for hematology
and blood bank testing,
which I'm sure you're very familiar with.
And then the next would be sodium fluoride
or potassium oxalate.
These tubes contain those
particular molecules
because they serve as
glycolytic inhibitors.
These basically preventing the
breakdown of of glu glucose.
And so these are gray top tubes
and we use them again as
I mentioned, typically
for glucose testing where
we wanna prevent any sort
of glycolysis that can occur.
And then finally the yellow
top tubes, these would be,
these are tubes that contain
an A, C, D, D solution,
either A or B, really
being for genetic testing.
That's what these tubes are used for.
And then there are some others
that come, you know, after that.
And it really is dependent
on institution if there's
research samples or things like that.
- Wow. Okay. I really appreciate this.
You kind of laid it out there.
And one of the things I
just want to go back to,
'cause I know it's often a
point of, of conversation,
is the difference between serum
and plasma.
Right? And you kind of
nicely highlight kind
of a stat version of this,
and this is what I love for our clinicians
to understand what's the
difference between serum plasma
and then for pathologists who
are in practice to think about
what are the timings
and what are the practicalities
for, for running.
So can you elaborate a little
bit on what is the difference
between serum plasma
and kind of why that serum
is gonna have that kind
of 30 minute additional
time tacked onto it?
- Sure. Yeah. So serum, as
I mentioned, that's going
to require clotting.
So that sample has to sit
for 30 minutes, clot has
to be generated, you
need to separate that.
And that is what's used for,
for a lot of our testing.
As I mentioned, that plasma sample, that's
what we use in our stat labs.
So again, doesn't need any
sort of clotting to occur.
I will say that there are
some differences in the
quality of the specimens.
Serum is typically a much cleaner specimen
compared to plasma.
So we try to use that when we can.
But understanding turnaround
team time needs, we will all
of the testing performed in the hospital,
clinical laboratory uses plasma
and we typ, we use the plasma
separator tubes or the PSTs.
- Wow, that's awesome to
understand that, you know, there,
there's pluses and minuses, right?
I think as you're kinda
laying that out, you know,
a cleaner sample with serum,
but it's gonna take longer to
kind of produce that serum.
In other words, for the
coagulation to go down and,
and kind of become devoid
of, of clotting factors
and fibrin in your sample
as opposed to, Hey,
I need it right now.
There's not time for that.
- Right. And ultimately, I
mean, it really comes down
to the specimen that's
chosen is what's acceptable
by the manufacturer for the
tests that are performed.
Some tests can only be
performed using plasma,
some are only validated for serum.
So we really have to, that's
kind of what's guiding
what tubes are collected.
- That idea, that concept.
I, I know there's probably
a decent percentage
of our audience that's
familiar with like, you know,
what is, is an approved sample
type for a particular test,
but maybe for our clinical audience and,
and our student audience
that's listening, you know,
what do we mean by that is, you know,
I think sometimes people
struggle with, you know,
here's the patient's blood.
Like why can't we test for this analyte?
- Yeah. So it really comes down
to the two different flavors of tests.
I'll, I'll say, so we
have FDA approved tests.
These have gone through the
rigors of being reviewed
by the FDA, the
manufacturers submitted it,
and there are specific claims in
what we call the package
insert that states it needs
to be tested using this specimen type.
It can be tested only on
this patient population.
These are the values
that you would expect.
These are the values that can be reported.
I know this often comes up, why
can't I have a lower result?
While that's not the, the,
the manufacturer has
not validated that, and
therefore, if it's not included
in their package insert,
we would be basically
going off-label, similar to
how you would be prescribing
a drug off-label for, for,
you know, when it's used for
its non intended purpose.
Hmm. So then the next would
be the lab developed tests.
And those are ones that,
you know, we've often
for esoteric measure,
for esoteric analytes,
but we, we measure it using,
let's say a mass spec or,
or some other instrument.
But that test is developed with parameters
that are within the lab.
So developed by the laboratory,
validated by the laboratory.
And I know there's a lot
of regulatory discussion
around this right now, and
there are changes coming,
but for all intents
and purposes from a
stat lab, we like to use
FDA approved tests.
So these are kits, you load
reagents on an instrument
and that testing is performed
according to the specs by the,
by the manufacturer.
- Hmm. Right. So you, those
are the ones where I guess
because it's been validated
that way, you, you, you're,
you're I guess, confident
that there's not gonna
be any interference.
- Yeah. So that, but,
and those interferences
are often listed in the
manufacturer's package insert.
So one of the interference that we deal
with a lot is hemolysis.
So the presence of free hemoglobin,
this can cause an interference
with the wavelengths
that are used for some of the tests,
or it just, you're dumping
contents out of the cell and
therefore it's not really a
true measurement of, you know,
that particular analyte.
So those things are spelled out and,
and we still, I i I
just wanna clarify this.
We still have to validate
that manufacturer's claim.
So the minimum requirement
we do is called par precision
accuracy, reportable range,
and then reference interval.
Those are the normal values.
So that is kind of the bare
minimum that we have to do.
We often do far more than
that as, as you can imagine.
But those are the requirements
to still show can we get the same values
that the manufacturer claims?
And, and we check this
for things like precision
manufacturer claims,
we can go down to this value
with this amount of precision.
And so we wanna know that,
that, that is the same
situation on our instruments in our hands.
- Well, you got me kind of
going down this pathway.
I'm really nerdy and curious.
I'm gonna have to invite you
back again to talk about,
you know, analytes and interferences. But
- That's my, that's my,
that's where my heart lies.
Pre-analytic interferences
are my favorite.
If you can, if there's a favorite
part of the testing phase,
pre-analytic is definitely
mine. I've spent a lot of
- My, I feel like academic
- Career
investigating interferences.
- Well, I feel that we'll
definitely bring you back
to come back to kind of the,
the order of collection.
And you brought up, you know, when errors
and mistakes are made, are
there ways that we can kind
of catch these errors?
Are, I mean, I guess
once it's done, I mean,
if something puts somebody puts
it in a different tube, it's
nearly impossible to detect.
Is that, is that right?
Or do we have ways of we
- Yeah, we do have some tricks
up our sleeves, fortunately.
So I'll just say in general, you know,
teaching toward the process
of correct order of draw is,
is just critical.
You know, we spend a lot of time on this.
We have our trainers and
trainees in a one-to-one fashion.
So they're training one-to-one,
and really they can't
perform any patient testing
until they've proven
that they can, you know,
operate using our standard
operating procedures without
any sort of assistance.
So that type of coaching
and mentoring really does
establish the muscle memory.
So the, you know, repeating it over
and over, it kind of becomes
ingrained in your mind,
this is not something that I do every day.
And so therefore, you know,
I am probably more rusty
than, than our phlebotomist.
But I think that is, is kind of critical
to setting the stage or,
or trying to prevent this from
happening in the first place.
And we do, you know, we
also do direct observations
and things like that to be
able to catch issues where,
where people may be making
these types of mistakes,
but from a lab perspective, we can co,
we can identify these
incorrect collections
using some creative ways to do this.
So we kind
of leverage the inaccurate
results that are produced.
So if a specimen is
collected inaccurately,
we use those inaccurate results
to help tip us off to say,
Hey, this probably wasn't
collected in the correct order.
So an example I'll give
for you is potassium, EDTA.
So EDTA is a molecule,
it's sequesters, divalent
and trivalent metal ions.
And so it's possible
that EDTA contamination
could be identified
by observing a decrease in
the concentration of some
of our commonly measured cation.
So that would be magnesium and calcium,
but we can also see large
elevations in potassium.
So I mentioned the tube
additive is potassium, EDTA,
and so we'll see very, very
high potassium concentrations.
And, and all of these components are
not compatible with life.
We, we, we can recognize that.
And what we use is our, our
middleware on the instruments
to flag those samples
and say, Hey, you might
wanna take a look at this.
And, and it's based off of
those patterns, a low magnesium,
a low calcium in the presence
of a high potassium up,
we're thinking EDTA contamination.
And so in that case, we
can reject that specimen,
prevent any results from being reported,
and we can recollect that specimen.
- That's awesome. And that's,
and you're running a STAT lab
and so even in the high
pressure environment,
this is functions well,
- Correct.
Yes. We, we rely pretty heavily on some
of those middleware rules
to help help us and guide us
and detect some issues with specimens.
- And I guess last question
I wanted to ask, you know,
are there some best practices for kind
of reinforcing this idea of
order of draw in the training?
Or is it just one of these
things of nothing to it,
but to do it and repeat, repeat, repeat?
- Yeah, I mean, that helps.
Repetition, as I mentioned, helps,
but I think the first thing I'll say,
and this is true of not
just laboratory collections,
but laboratory testing in
general, really helping
to un have our techs
understand the why behind it,
behind it, why is it so important?
And I think sharing, you know,
what would happen if you
didn't do this correctly?
And then translating that
into the patient impact all
of our staff, they wanna
do the right thing.
They wanna make sure that they're giving,
they're providing samples
that provide the best
results for our patients.
And so I think it's really helpful to,
to help them understand
that their work is not only valuable,
but the way they do it is
critical for accurate testing.
So that in general, I
think is probably the best,
the best way to address order of draw and,
and maintain, you know,
appropriate procedures for that.
The next, I I sort of mentioned this,
but regular observations, you know,
more frequently assessing
what people are doing.
It helps identify that problem,
but it also helps us identify other
opportunities for improvement.
So not just the order of draw process,
but all facets of collections.
It helps give us an idea of
what may be problem spots,
what may be issues in our
SOPs that we need to address.
So I think doing that along
with positive reinforcement
is really, you know, a useful tool
to help encourage the desired behavior.
And, and this is happening,
you know, we often, we,
we do our annual competency,
but doing those direct
observations more frequently,
monthly, for example, allows
you to catch any issues,
you know, a bit earlier than you would
with just an annual competency review.
- I can imagine that that also kind
of normalizes the experience
of being observed, right?
Correct. If it's happening
more frequently, yes.
Rather than like, oh,
here's my observation,
so now I'm really gonna
do things correctly.
If it gets more exactly into
the general habit, you know,
maybe that makes it a little bit more
of a welcoming environment
and also maybe puts me just
doing my normal routine.
- Right. Right. And I'll
say the, I mean the,
the last thing I'll offer is from the
laboratory perspective.
So, you know, if we detect these errors
or we suspect them, it's
really important that we share
that back with the phlebotomy team
and particularly in a timely fashion.
This kind of goes back to
the direct observations.
If we can do this more
quickly, we can, you know,
nip it in the bud so to speak,
and try to adjust these issues.
Maybe it's something that a,
that a phlebotomist is just,
they just aren't unsure about the
process and they keep doing.
And so if we can kind of
catch that and prevent it,
and we accomplish this
through using our lab event
management system, so lems,
that allows the phlebotomy
team to perform, you know,
targeted education and address any issues
or behaviors as they
occur so that we can try
to prevent these from happening.
And, and we do a great job at this.
Every once in a while we have,
we kind of have these flags
and say up looks like EDTA contamination,
but in general we do a great
job of managing that and,
and making sure that we
are doing it correctly.
- Wow. We've been rounding with Dr.
Brooke Katzman, understanding
the why behind the Rainbow
of Tube collection.
Thanks for joining us Dr. Katzman.
- Thank you so much. Thanks for having me.
- If you've been interested in registering
for Mayo Clinic Laboratories
upcoming phlebotomy conference,
please check out the the show notes.
And thank you for taking
the time to talk with us.
To all of our listeners, thank
you for joining us today.
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