Hi, I'm Bobbi Pritt, Director of the Clinical Parasitology Laboratory and vice chair of education in the Department
of Laboratory Medicine and Pathology at Mayo Clinic.
In this month's Hot Topic, my colleague Dr. Brad Karon will discuss the need and evidence behind
following the order of draw recommendations for routine blood collection, specifically addressing whether
the existing evidence demonstrates a need to collect serum tubes before either potassium EDTA or citrate
tubes.
I hope you enjoy this month's Hot Topic and I want to personally thank you for allowing Mayo Clinic
the opportunity to be a partner in your patients' healthcare.
Thank you, Dr. Pritt.
To those of you who have not attended our phlebotomy conference in the past, the title of today’s
session, “Phlebotomy Top Gun,” may seem a little odd, I hope I’ll be able to explain
to you what is “Phlebotomy Top Gun” all about and what we try to do at our annual
phlebotomy conference.
I have no disclosures relevant to today’s presentation.
Every year at our phlebotomy conference, I present a talk that’s always called, “Phlebotomy
Top Gun.”
The format for “Phlebotomy Top Gun” is a case-based presentation.
I solicit from attendees of the conference, the questions, issues, areas of concern they
would like to hear about.
Using the questions submitted by the audience, I present these as case-based scenarios, using
an audience response voting mechanism, the attendees of the conference vote on what they
believe the correct answer to the case or question to be.
I then present the collective experience of the Mayo practice, as well as any published
data and evidence I can find related to the topic.
At the end of the case presentation, the audience votes again.
For each case, I can see whether I’ve been able to change anyone’s mind on the question
at issue by presenting the data, evidence, and information relevant to this particular
topic.
This is an actual case from a last year’s “Phlebotomy Top Gun” presentation.
In last year’s presentation we were addressing the issue of order of draw and whether the
order of draw is still relevant in today’s modern phlebotomy practice.
So the question I posed based on a question submitted by a conference attendee last year
was the following:
Which of the following statements describes the importance of the drawing a serum tube
before the K EDTA tube?
In the era of lyophilized anticoagulants, order of draw is no longer important.
Contamination of serum samples with K EDTA will occur routinely if order of draw is not
followed.
During syringe collections contamination with K EDTA may rarely occur if order of draw is
not followed.
Modern lab equipment can tell the difference between K from EDTA tube and K from the patient
so order of draw is no longer important.
At this point in the phlebotomy conference attendees will vote on what they believe to
be the right answer at the end of the case we’ll re-poll and I’ll reveal what I believe
to be the right answer and I’ll get to see if I changed anyone’s mind.
A great number but not all cases start with a review of the CLSI guidelines which have
a great deal of information about recommended procedures for blood collection.
The relevant guideline in this case which is often the case, is the GP41, collection
of diagnostic venous blood specimens guideline from the CLSI, in the guidelines they go over
order of draw recommendations which continue to be present in the guidelines as blood culture
first, followed by citrated tubes, serum, heparin, then EDTA with special emphasis on
not collecting the EDTA tube before the serum tube.
The guidelines do clarify that although these recommendations are valid for both liquid
anticoagulants, glass tubes, which are really not used other than a few special coagulation
type tests in our practice, and the more commonly used plastic tubes that have lyophilized additive
in them.
The guidelines go on to say for additives not listed that you consider the type of additive
and where it fits in to the order of draw.
The guidelines suggest not collecting serum clot activator tubes before a citrated tube
for coagulation study to avoid carry over of the clot activator into the coagulation
tube.
They specifically mention the guidelines that Na carry-over from Na citrate or Na heparin
tubes into a serum tube is not really significant and then for trace metals follow the manufacturer’s
instructions for various tubes and collection devices.
That’s a review of the CLSI guidelines, again where I start with many of the cases
presented or questions presented at the phlebotomy conference we start reviewing CLSI guidelines.
In the case of order of draw this is one of the rare situations where we have a CLSI guideline
to consider, we also have this European Federation for Clinical Chemistry Working Group on pre-analytical
phases of testing that really did an extensive review and I have references at the end of
this presentation for those of you who would like to read it, but this European group really
did an extensive review and had an consensus conference on a number of pre-analytical phases
or pre-analytical questions about blood collection.
This document goes on to list various problems that have been linked to incorrect order of
blood draw, including hypernatremia due to Na citrate or Na EDTA, which if you recall
a slide ago CLSI said it was not an issue, the European Federation suggested that it
could be an issue.
Hyperkalemia due to K EDTA contamination, that’s really what we are going to focus
on, is K contamination from K EDTA tubes into serum.
Hypocalcemia, low MG, zinc, alkaline phosphatase, other enzymes again due to EDTA contamination
of serum tubes.
Poor coagulation due to the transfer of anticoagulants, clot activator for example into citrated tubes,
clot activator interference with coagulation testing as I mentioned and dilution effects
if you pour one tube into another.
These guidelines then went beyond what CLSI has done which provides you guidelines for
what you should do in a few references, this again, I encourage people to read this, a
very nice document to go into why this is important or basically what are the mechanisms
of K EDTA contamination and how much evidence exists that they do or don’t actually happen.
They went over three potential mechanisms for anticoagulant contamination for K EDTA
contamination of serum tubes when the order of draw is not followed.
Direct transfer or pour off, again obviously if you need serum, you collect K EDTA, you
pour that into a serum tube, you’re going to have contaminated sample, obviously a very
bad practice and we don’t expect to see that where we have trained phlebotomists.
Backflow during closed loop (vacutainer) collection, so if you snap the K EDTA tube on to the vacutainer
adaptor there is potential for some blood that has K EDTA on it to stick to the vacutainer
adaptor and that could get transferred in to the serum tube.
We’re going to show you in a minute that the studies really don’t suggest that actually
happens with lyophilized K EDTA, so this is what may have changed going to plastic tubes
with a lyophilized reagent, like lyophilized anticoagulant we don’t see that there is
carry-over from backflow on a vacutainer adaptor.
And then contamination by syringe, so if you use a syringe to dose the tubes in the incorrect
order, so dosing the K EDTA first then the serum there is the potential to carry-over
some of that K EDTA from the purple top K EDTA tube to the serum tube.
Data on this, weather is really happens and how much often this happens is somewhat spotty
or controversial but really that is what we are going to talk about, if this doesn’t
happen anymore then maybe order of draw is no longer important.
However, in general, what the European Federation for Clinical Chemistry, this pre-analytical
work group concluded is, order of draw was not important for routine venipuncture using
a vacutainer adaptor but that contamination still did occur during syringe transfer and
because it did rather than training one way for routine vacutainer and another for syringe
it was easiest to follow order of draw as defined in CLSI guidelines and by this guideline
group all the time.
These studies are really those sited by the European Federation working group document.
One study of 11 healthy volunteers, drawn by experienced phlebotomist.
In this experiment they drew a serum tube, K EDTA tube, then a second serum tube to just
try to measure in a healthy outpatient using experienced phlebotomists, was there any measureable
EDTA carried over from the middle tube into the second serum tube.
Well the answer was no, EDTA was not detectable in any of the serum samples and looking at
the measurements that would be affected by K EDTA contamination.
K obviously, and then the metals, calcium, magnesium, zinc, and the enzymes that need
calcium and metals like alkaline phosphates, there was no difference between the serum
tube before the EDTA and the serum tube drawn after.
So it is a small study, 11 healthy volunteers, but really didn’t find that contamination
occurred when you drew an EDTA tube before the serum tube.
Little bit larger study here, sited 57 outpatients attending an anticoagulation clinic and 58
healthy volunteers now we’re getting to 120 or so patients involved, multiple experienced
phlebotomists.
Again outpatients, volunteers, drew a serum then a citrate then a second serum and the
other set of volunteer serum, K EDTA, then a serum.
And again similar results, there is no difference between the mean K, Na, calcium, MG, phosphorus
in the serum collected first and the serum collected after and EDTA or Na citrate tube.
Consistent with the smaller study I just showed, really showing that order of draw probably
doesn’t matter in routine outpatient phlebotomy using a vacutainer adaptor were not seeing
carryover between tubes.
In contrast here’s a study done on hospital inpatients in one UK hospital and for one
month they simply said, well let’s look at all inpatient blood serum K, where the
K is over 6 mmol/L and lets just see if we can detect measurable EDTA in those.
So they had 117 samples with serum K over 6 mmol and 25% of those had detectable EDTA,
over 0.1 mmol/L in this study.
So that would suggest that carryover is fairly frequent, just selecting serum samples with
high K, we’re seeing a lot of these samples, almost a quarter of them, have some amount
of EDTA that presumably had to come from a K EDTA tube, there is not another logical
explanation for where EDTA in a serum sample would come from.
Again, most of these were inpatients and almost all of these patients when they had their
K measured a second time that started over 6 returned normal and had no EDTA so again
further evidence that this was likely K EDTA contamination in these patients with high
K serum values.
EDTA concentration all were measurable but the more EDTA the higher the K, the lower
the calcium, zinc and MG and alk phos, so more evidence that what their seeing is a
variable amount of K EDTA getting into serum samples somehow.
The study didn’t say how it got there just showed it got there and that it caused high
K, low calcium, MG, zinc and alk phos.
The same group of authors went on to do a multi-center study to say well this was this
one UK hospital let’s look at five hospitals, these hospital were all in the UK as well,
but looked at five different hospitals over a month, again very similar idea, let’s
look at all serum K over 6 and let’s see how many of those had measurable EDTA to see
if this was a fluke.
You can see they looked at different types tubes, Sarstedt vs. Greiner, and one site
had BD.
Each site had between 100 and 300 of these serum samples with K over 6 and then you can
just look at the last column, the percent that had EDTA contamination varied from 1%
to almost 7%.
That very first study showed almost 25% of these serum samples had K over 6 contaminated
with EDTA, here we’re seeing it still happens but lower numbers of 1-7% of these serum samples
with high K are contaminated by K EDTA.
What’s our experience been like here at the Mayo Clinic?
We also do a similar thing, we have a number of rules built into our middleware system
in the lab to detect possible contamination with either K EDTA or IV fluid, but specifically
there is a flag that goes if the K is over 6 mmol on an inpatient and then it will look
at other values that are requested with that K and alert for potential contaminations.
When the flag goes off for potential contamination a caregiver is contacted.
If this value is not expected for this patient, we’ll redraw it and most of the time we
find that the K comes back relatively normal if it’s a contamination suspected.
Most of the time that K contamination is from K given intravenously to the patient not suspected
to be K EDTA.
Unusual cases that we reviewed, go to a resident or fellow on-call and we review those weekly
in our call rounds and it seems like once a quarter or once a month we have a sample
with a very high K that doesn’t look real and often times a resident or fellow will
say if they didn’t order a calcium or MG can we measure one to see and it will come
back similar to the studies that I showed with a very high K, a very low calcium or
MG.
It will be redrawn with a normal K and it will look like K EDTA contamination similar
to the studies I showed.
We don’t exactly know, we don’t go back to know if it was drawn with a syringe or
a vacutainer we suspect what we’re seeing here is what’s been shown in other studies,
some low rate, 1%, 5%, of these very high serum K samples have somehow been contaminated
K EDTA, we suspect during syringe draws that were done improper order of draw was followed
so the K EDTA tube was dosed first and then the serum tube.
Conclusions for order of draw: evidence suggests that K EDTA contamination does not occur when
you have a closed draw that is a vacutainer needle and vacutainer adaptors with plastic
tubes and lyophilized K EDTA it doesn’t really look like there’s backflow is still
happening that K EDTA gets stuck to the vacutainer adaptor and goes into the serum tube if its
drawn after an EDTA tube for a vacutainer collection.
The greatest data on syringe draws comes from these five UK hospitals, the study I showed,
K EDTA contamination is seen in somewhere between 2-15% of all samples that had very
unusual K, calcium, MG or zinc concentrations.
I want to make clear, it’s not that 2-15% of all serum samples are contaminated with
K EDTA, but when you select for something like K over 6 or very low calcium or MG values
then you see the 2-15% of those samples do appear to be contaminated K EDTA.
So it seems that order of draw is still important and with syringe draws particularly you can
carry over K EDTA from the purple top to the serum tube if you do not dose them in the
correct order or essentially follow the recommended order of draw.
At Mayo Clinic we are still rarely seeing samples that look like they have K EDTA contamination,
in our practice we were able to figure out is that a vacutainer or syringe collection.
It could be happening more often then we’re detecting with our middleware rules and systems.
The underlying assumption I’m left with is that order of draw is still important;
you should draw the serum before the K EDTA purple top tube and the underlying assumption
from the data and evidence out there, contamination happens primarily with syringe draws, it doesn’t
seem that backflow contamination with a routine vacutainer collection, but I would agree with
the European Federation Workgroup guidelines, if you going to do it for syringe draws its
probably just easiest to follow the recommended order of draw for all blood draws.
During the phlebotomy conference we would then go back and re-poll the audience, now
that we have given our experience and reviewed the published data and guidelines that are
out there and we would go back and now re-poll our audience the same question.
Which of the following statements describes the importance of the drawing a serum tube
before the K EDTA tube?
Again, the same answers: In the era of lyophilized anticoagulants,
order of draw is no longer important.
Contamination of serum samples with K EDTA will occur routinely if order of draw is not
followed.
During syringe collections contamination with K EDTA may rarely occur if order of draw is
not followed.
Modern lab equipment can tell the difference between K from EDTA tube and K from the patient
so order of draw is not important.
The first three of these are plausible, the fourth I just threw in for fun, no our lab
equipment can’t tell the difference, K is K and we would allow the audience to re-poll
and I would present what I believe to be the answer, which is number three, that order
of draw is still important because during syringe collections contaminations with K
EDTA may rarely occur if the order of draw is not followed.
So that’s the end of my presentation and again this is case one of four or five I presented
at last year’s “Top Gun Phlebotomy” presentation at our phlebotomy conference,
if you enjoyed this presentation, or even if you didn’t, I would encourage you to
investigate, register for and come to our Phlebotomy 2019 Conference: People + Performance
= Patient Care, more information can be found on the Mayo Clinic Laboratories website.