Quick on the draw—coagulation tube response
As the sensitivity of coagulation testing has increased,
the preanalytical phase has been getting more attention as a potential source
Variables that have long been known to affect the accuracy of activated partial
thromboplastin times are the reagents and instruments used in testing and the
delays between acquiring and processing a blood sample.
More recently, the coagulation tube itself has been found to be a clinically
significant source of variability. So significant, in fact, that it led the
largest supplier of sodium citrate tubes worldwide to withdraw a line of tubes
from the market in September 2000.
Becton Dickinson, Franklin Lakes, NJ, stopped manufacturing its partial-draw
sodium citrate tubes after several clinical studies found APTT times were artificially
shortened in patients on unfractionated heparin. Platelet counts also tended
to be falsely low.
"Partial-draw tubes were introduced into the market because people wanted
to draw a lower blood volume, but everything had to be standardized around a
certain tube size, 13 x 75 mm," says Jeffry B. Lawrence, MD, vice president
for medical and scientific affairs, BD Clinical Laboratory Solutions. "The
idea was you would have the best of both worlds: You would be drawing less blood
but you would also be able to use your same instruments and centrifuge and blood
This simple design meant that the 13 x 75 mm glass tube contained significantly
less blood than the standard full-draw tube. The partial-draw tubes were manufactured
with less vacuum, resulting in 1.8-mL or 2.7-mL blood draws.
"At the time we withdrew the product, we got our engineers together to
fix the problem because the marketplace needed tubes in the 13 x 75 mm configuration,
but we couldn’t use the partial-draw design because it caused erroneous results,"
says Dr. Lawrence. "The lower blood volume in the tube was associated with
more headspace—the vertical distance between the top of the column of
blood and the stopper.
"Partial-draw tubes fill more slowly than full-draw tubes," he adds,
"and when these tubes are filled, the platelets get activated by prolonged
exposure to shear forces arising from drawing blood into the increased headspace.
The activated platelets then release platelet factor 4, which neutralizes heparin
in the samples." This results in a lower effective concentration of heparin
in the citrate tube than is present in the patient, which artificially shortens
the APTT in patients on heparin therapy.
BD is also eager to "get out of glass wherever clinically feasible,"
says Dr. Lawrence. "We were increasingly concerned about the risk of health
care worker injury from the breakage of glass tubes."
In May, the company launched a new line of low-volume, plastic coagulation tubes
that are being marketed as BD Vacutainer Plus plastic citrate tubes. They consist
of a tube within a tube. The outer tube is made of polyethylene terephthalate
and has standard 13 x 75 mm external dimensions. By varying the diameter of
the polypropylene inner tube, a minimized headspace is maintained in both the
1.8-mL and 2.7-mL draw volumes.
"We studied more than 1,000 patients with these new Plus plastic tubes,"
says Dr. Lawrence. "We didn’t approve their release until we showed that
across all patient populations there wasn’t any effect on the test results."
People have long undergone coag ulation testing for clinical
indications such as abnormal bleeding and excessive clotting and for screening
prior to eye, brain, and other major surgeries in which excessive blood loss
could cause problems. More recently, labs that perform coagulation testing have
been focusing on monitoring patients on anticoagulation therapy, either intravenous
heparin or oral warfarin.
"One of the reasons that this has become more important from a clinical
perspective," says Dr. Lawrence, "is that people are using a lot more
anticoagulants. It’s been shown that you can really reduce the morbidity and
mortality rate from heart attacks and from certain other kinds of clotting problems
if you anticoagulate these patients."
Because it’s impossible to predict the anticoagulant dose needed to treat a
patient, that person is typically given a dose of intravenous heparin, for example,
and six hours later an APTT is performed to see if it falls within a certain
therapeutic range. "The reason why it is important to standardize tube
performance is that the therapeutic ranges of heparin and warfarin are the best
compromise between a lower dose that may not prevent clotting and a higher dose
that may cause bleeding," Dr. Lawrence says.
A published review of several clinical trials has found that four percent of
patients on intravenous heparin therapy for thrombotic disease will have major
bleeding as a consequence. "No one knows what number occurs because of
inaccurate coagulation results, but I think a significant number of them could
be due to issues such as the partial-draw tubes," Dr. Lawrence says.
BD sponsored clinical studies in laboratories nationwide to test the performance
of its new tubes on healthy subjects and patients with coagulation disorders
on anticoagulant therapy. The assays performed were anti-factor Xa heparin,
APTT, fibrinogen, platelet counts, and prothrombin time.
In each case, the performance of the new 1.8-mL and 2.7-mL low-volume plastic
tubes was compared to the full-draw, 4.5-mL glass tube. "We refer to that
as the gold standard," says Dr. Lawrence, "because the only information
that exists about what dose a patient should have based on their PT or APTT
[comes from studies] done with BD full-draw glass tubes."
Furthermore, because the degree of bias observed with the partial-draw tubes
varied depending on the reagents and instruments used by a laboratory, the tubes
were tested with the following reagents: Dade Actin (FS, FSL), Dade Innovin,
Dade Thromboplastin C Plus, Hemoliance Brain Thromboplastin, Hemoliance Thrombosil
1, IL Test APTT-SP, IL Test PT-Fibrinogen HS Plus, Stachrom Heparin Xa reagent,
and STA Fibrinogen. The tubes were also tested with the following instruments:
Coulter STKS, Dade Behring BCS, Diagnostica Stago STA, IL Electra 1400C, IL
Futura, Sysmex 1500C, and Sysmex K1000. "We tried to look at the market
share in and outside the United States to find the most prevalent reagents and
common instruments being used," Dr. Lawrence says.
Julie Wengert, MT(ASCP), a blood bank/hematology supervisor at Southeast Missouri
Hospital, Cape Girardeau, Mo., was involved in testing BD’s partial-draw tubes
before they were withdrawn from the market and in testing the new low-volume
"Basically, their tube is a tube in a tube. They made them so that the
tubes are completely full, there is no dead space, and there is no platelet
activation," she says. The studies in Wengert’s laboratory found that the
APTTs and PTs obtained with the new 1.8-mL and 2.7-mL tubes were comparable
to those obtained with the full-draw 4.5-mL tube.
The results paralleled those from another site where BD sponsored studies of
the new tubes: No remarkable difference between them and the full-draw tube
was found, says David R. Chance, BS, SH(ASCP), technical supervisor at the Saint
Louis (Mo.) University Hospital Coagulation Reference Laboratory. Furthermore,
he adds, "There is the safety factor of not having to worry about broken
Another advantage of plastic is lower disposal costs, says Norman Anderson,
MT(ASCP), laboratory director at Southeast Missouri Hospital. "When you
look at glass versus plastic and disposal and waste, you’re looking at volume,
and [plastic] would really help costs because glass weighs more."
Dr. Lawrence believes the clinical trials sponsored by BD can satisfy the Clinical
Laboratory Improvement Amendments of 1988 as well as CAP requirements for documenting
the performance of the various tools used in the lab.
"Under the CLIA ’88 legislation, the laboratory director has responsibility
and liability for all clinical diagnostic testing in their laboratory,"
he says. "If I introduce something in my lab that hasn’t been adequately
clinically validated, I could be at risk. Both for CLIA compliance and CAP inspections,
they [labs] need to have on file documents that show whatever product they’re
using does not increase any artifacts."
While this information can be provided by the medical literature or independent
laboratory studies, in Dr. Lawrence’s experience, it’s often provided, or at
least augmented, by the manufacturer.
"Most laboratories are extremely short-staffed and it’s not cost-effective
for them to do their own extensive studies," he says. "By and large,
customers find it extremely helpful for us to provide them with the papers."
In contrast, the No. 2 supplier of coagulation tubes worldwide, Greiner Bio-One,
Monroe, NC, has found its customers prefer to do their own studies. "’I
have to prove it to myself in my own laboratory,’" is what Doug Harris,
vice president of marketing and sales, says he hears repeatedly from customers.
"At the same time," he says, "most laboratories also want to
examine their ranges to ensure that they are appropriate with the particular
coagulation test system employed."
Greiner’s Vacuette sandwich coagulation tubes have been on the market since
1996. Like BD’s tubes, they have an inner tube made of polypropylene and an
outer tube made of polyethylene terephthalate. The outer tube has the standard
13 x 75 mm outer dimensions, and the inner tube has a 9.5-mm diameter.
In the United States, Greiner manufactures three draw volumes: 1.8 mL, 2.7 mL,
and 3.15 mL. "Our 2.7-mL tube has less headspace than the original 4.5-mL
tube, and the 1.8-mL tube is similar as well," Harris says. "Furthermore,
we have not yet shown in studies we’ve formally conducted that any of our tubes
produce changes that could be clinically significant."
Greiner is not convinced that headspace is primarily responsible for the phenomena
exhibited with partial-draw tubes. "It was not a consistent finding in
every tube," Harris says. "No one could exactly say what was causing
the problem because coagulation is a very complex science." Instead, Greiner’s
preanalytical focus has been on activating platelets by glass and the blood-drawing
process. Other factors, he says, are the variability between reagents and instruments.
Greiner has started manufacturing its coagulation tubes with harder stoppers.
It made this change to improve the stoppers’ response to automated cap-piercing
Greiner’s tubes also have a nominal fill line, which is possible with plastic
because, unlike glass, it’s a molded, not extruded, product. This reference
line indicates that the tube is filled properly and helps to avoid variability
in the blood draw volume, which is influenced by the tube’s shelf life, patient
blood pressure, angle of vein entry, and other factors. "It’s important
that the tube is always filled correctly so that a 1-to-9 citrate-to-blood ratio
is achieved," Harris says.
Though many laboratorians were frustrated when BD pulled
its partial-draw tubes off the market two years ago, Dr. Lawrence says, "It
was an easy decision for us to make. This new product requires no compromise:
It give health care workers the safety of plastic; it gives documented clinical
performance; and it draws a lower volume of blood without the partial-draw configuration."
Wengert and Anderson, too, believe BD acted appropriately. "They address
problems," says Anderson. "From my experience as a laboratory manager,
I respect vendors who address problems rapidly so we can rapidly respond, and
I know that costs the vendor money."
Vida Foubister is a writer in Mamaroneck, NY.