Case Report

Use of therapeutic plasmapheresis in a case ofcanine immune-mediated hemolytic anemiaKathryn L. Crump, BVSc and Ravi Seshadri, DVM, DACVECC, DABVP

Abstract

Objective To investigate the clinical application and potential utility of plasmapheresis in canine immune-mediated hemolytic anemia.

Case Summary A 7-year-old spayed female Maltese diagnosed with immune-mediated hemolytic anemiawas initially treated with prednisone, cyclosporine, and received multiple transfusions of packed RBC.Because of the progression of clinical signs despite traditional medical therapy, plasmapheresis was initiated.Plasma immunoglobulin G and immunoglobulin M levels were measured before, during, and after treatmentto help determine if there had been a significant decrease in immunoglobulin levels with plasmapheresis.Plasmapheresis was successfully performed over a 2.5-hour period in this dog with minimal complications.Hypocalcemia was identified as a known complication of circuit anticoagulation, and was corrected throughcalcium supplementation. Post-plasmapheresis there was a decrease in immunoglobulin G and immunoglobulinM levels, and the patient showed clinical improvement. Following discharge the dog had no knowncomplications of therapy, and had complete resolution of the anemia.

New or Unique Information Provided Plasmapheresis was performed successfully with minimalcomplications. Because transfusion requirements appeared to be reduced, and the procedure was welltolerated, there may be a place for this modality in severe cases to act as a bridge until medical therapy takesfull effect. Because of the cost of performing this therapy, and the potential requirement for multipletreatments, it should be reserved for selected patients.

(J Vet Emerg Crit Care 2009; 19(4): 375380) doi: 10.1111/j.1476-4431.2009.00431.x

Keywords: autoimmune, hemolysis, plasma exchange, RBC, transfusion medicine

Introduction

Therapeutic plasmapheresis was first used to treat

hyperviscosity syndrome in humans and has been ap-

plied to diseases ranging from autoimmune disorders

to toxicities.1 Plasmapheresis reduces the levels of an-

tibodies in the plasma and if effective, leads to a tem-porary remission of the disease.2,3 After separation of

plasma from the whole blood, the plasma can be treated

and returned to the patient, or discarded and replaced.

In human immune-mediated disease, plasmapheresis is

commonly used in combination with immunosuppres-

sive protocols to achieve rapid control of clinical signs,

and to reduce adverse effects of therapy. In veterinary

medicine, the use of plasmapheresis has been reportedfor systemic lupus erythematosus,4 immune-mediated

hemolytic anemia (IMHA),5 myasthenia gravis,6 and

hyperviscosity syndrome.7 Previous veterinary reports

have used centrifugal cell separation technique. The

case described here is the first known veterinary report

for IMHA utilizing membrane filtration technique.

One of the most common immune-mediated diseases

seen in dogs is IMHA. Treatment traditionally involves

immunosuppression with steroids in combinationwith cyclosporine, azathioprine, IV immunoglobulin,

or other immunosuppressive medications.2,8,9 The

pathophysiology and treatments for IMHA have been

well described in the veterinary literature, with

reported treatment success rates of 4070%.8,10 For

refractory cases treatment options can be limited, and

animals may require multiple transfusions before med-

ical therapy takes effect to slow autoantibody produc-tion and hemolysis. The case described in this report

was managed with a combination of traditional immu-

nosuppressive therapy, and a single plasma exchange

for treatment of IMHA. Immunoglobulin levels were

quantified to help monitor the efficacy of clearance of

immunoglobulin G (IgG) and immunoglobulin M (IgM)

with treatment.

Address correspondence and reprint requests toKathryn Crump, 1409 Clark Lane, Redondo Beach, CA 90278, USA.Email: kathcrump@gmail.com

From the Advanced Critical Care and Internal Medicine, Tustin, CA 92780.

Journal of Veterinary Emergencyand Critical Care 19(4) 2009, pp 375380doi:10.1111/j.1476-4431.2009.00431.x

& Veterinary Emergency and Critical Care Society 2009 375

Case Summary

A 7-year-old spayed female Maltese, weighing 4 kg,

was referred to Advanced Critical Care and Internal

Medicine for further evaluation of acute anemia, leth-

argy, and anorexia. Bloodwork at the primary veteri-

narian showed a PCV of 17% (reference interval3755%), total plasma protein (TPP) of 62g/L (6.2g/dL)

(reference interval 5878 g/L [5.87.8 g/dL]), platelet

count of 737 109/L (reference interval 175500 109/L), bilirubin of 17.1mmol/L (1.0mg/dL) (refer-

ence interval 015.4mmol/L [00.9mg/dL]), and alka-

line phosphatase of 631 IU/L (reference interval

5131 IU/L). Remaining parameters were within refer-

ence intervals.Physical examination abnormalities included pale

mucous membranes and mild clinical dehydration.

Other than mild hepatomegaly, the abdominal and tho-

racic radiographs were within reference limits. The

PCV was 18% with TPP of 72 g/L (7.2 g/dL). A saline

autoagglutination test was performed and was strongly

positive. Coagulation parameters were within reference

intervals with a prothrombin time of 9.5 seconds(reference interval 6.711.4 s), and activated partial

thromboplastin time of 14.1 seconds (reference interval

11.920 s). The dog was mildly tachycardic with a heart

rate (HR) of 140/min, and tachypneic with a respira-

tory rate (RR) of 55/min. The patient was transfused

because she was clinically affected by the anemia, with

tachycardia, tachypnea, and clinical weakness noted on

physical examination. Mild ptyalism was noted, andthe dog retched several times after abdominal palpa-

tion. A transfusion was started using 15mL/kg of

washed, leukocyte-reduced packed RBC that increased

the patients PCV to 30%. The TPP was 70 g/L (7.0 g/

dL). The dog appeared clinically brighter with a stable

HR, RR, and systolic blood pressure of 114mm Hg using

an ultrasonic Doppler flow monitor. An abdominal

ultrasonographic examination was within reference lim-its apart from mild hepatosplenomegaly. The dog was

started on prednisonea (1.8mg/kg, PO, q 12h), cyclos-

porineb (5mg/kg, PO, q 12h), sucralfatec (50mg/kg, PO,

q 8h), dolasetrond (0.6mg/kg, IV, q 24h), doxycyclinee

(5mg/kg, PO, q 12h), famotidinef (0.5mg/kg, PO, q

24h), enoxaparing (1mg/kg, SC, q 12h), and balanced

isotonic crystalloid fluidsh (60mL/kg/d).

The CBC submitted on initial presentation showed anormocytic, normochromic anemia with a HCT of 16%

(reference interval 3755%), WBC count of 15.6 109/L(reference interval 5.716.3 109/L) with a neutro-philia of 13.1 109/L (reference interval 3.011.5 109/L)and a lymphopenia of 0.94 109/L (reference interval1.04.8 109/L), moderate polychromasia, anisocyto-sis, and slight spherocytosis. The anemia was classified

as regenerative (absolute reticulocyte count 139 109/L; reference interval 060 109/L), and the Coombstest was positive at a dilution of 1:64. Over the next

3 days of hospitalization the PCV continued to rapidly

drop and 3 further transfusions of packed RBC were

required.

Options considered for treatment included oxyglo-bin, additional immunosuppressants, IV immunoglob-

ulin, and plasmapheresis. Plasmapheresis was chosen

to reduce the rate of hemolysis allowing more time for

immunosuppressive therapy to take effect. Informed

consent was obtained. The pretreatment PCV was 12%

and TPP 62g/L (6.2 g/dL). The patient was transfused

with 10mL/kg packed red cells before and during

plasmapheresis. The pretreatment temperature, HR, RR,and blood pressure were within reference intervals. CBC,

venous blood gas/electrolyte panel, serum biochemical

profile, and prothrombin time and partial thromboplastin

time were performed. Abnormalities included an

alkaline phosphatase of 295 IU/L (reference interval

5131 IU/L) and bilirubin 35.9mmol/L (2.1mg/dL)

(reference interval 015.4mmol/L [00.90mg/dL]). A

pretreatment sample was obtained and frozen at 181C(01F) for 2 weeks for immunoglobulin analysis.An 8-Fr double-lumen, short-term, silicon dialysis

catheteri was placed using the percutaneous Seldinger

technique. A continuous renal replacement therapy1

therapeutic plasma exchange machinej was used along

with a plasma filterk with an effective surface area of

0.35m2, and a total extracorporeal circuit volume of

88mL. Immediately before use, the blood access lines,hemofilter, and total extracorporeal circuit were primed

with 0.9% saline. Fresh frozen plasma was chosen as the

replacement solution. The plasma volume was calcu-

lated using the following formula:

Plasmavolume mL 1HCT total blood volumePlasmavolume mL 1 0:7 90 4 252 mL

The dogs plasma volume was estimated to be250mL (PCV was estimated at 30% as the patient was

receiving a transfusion during plasmapheresis). Plasma-

pheresis was performed by removing 500mL of plasma

from the patient, with concurrent replacement of

500mL of fresh frozen plasma delivered over 2.5 hours.

The blood flow rate through the extracorporeal circuit

was set at 20mL/min (using human guidelines of 2

5mL/kg/min). Sodium citrate anticoagulationl of theblood access line, blood return line, and plasmapheresis

filter was started using a commercial sodium citrate

solutionl at 18mL/h delivered through a Y-set at the

proximal blood access line. A calcium chloride

solutionm (56mg/h) was administered through the

peripheral catheter to counter hypocalcemia.

& Veterinary Emergency and Critical Care Society 2009, doi: 10.1111/j.1476-4431.2009.00431.x376

K.L. Crump & R. Seshadri

Plasmapheresis was well tolerated, with mild hypo-

thermia of 37.51C (99.51F) countered through IV fluidline warming and cage heat support. Hypocalcemia(ionized calcium, 0.93mmol/L; reference interval

1.121.4mmol/L) was noted within 30 minutes of start-

ing plasmapheresis, so the calcium chloridem infusion

rate was increased gradually to 70mg/h to counter this.

Throughout treatment the electrolyte/blood gas panel

was checked every 30 minutes with samples drawn

from the extracorporeal circuit and the patient. If the

extracorporeal circuit calcium was 40.4mmol/L thesodium citrate rate was increased to improve anticoag-

ulation of the blood in the circuit, and prevent the blood

filter from clotting during treatment. Additional serum

samples were obtained every 30 minutes and frozen at

181C (01F) for 2 weeks for immunoglobulin analysis(see Table 1).

Postplasmapheresis diagnostic testing showed a PCV

of 42% and TPP 50 g/L (5.0 g/dL). A CBC wasperformed and was within reference intervals. The pa-

tients acid-base status remained normal although ion-

ized hypocalcemia persisted (0.7mmol/L, reference

interval 1.121.4mmol/L). The chemistry panel

showed mild hypoalbuminemia (22 g/L [2.2 g/dL],

reference interval 2339 g/L [2.33.9 g/dL]). The hypo-

calcemia improved with the ionized calcium increasing

to 1.14mmol/L within 6 hours without the need forfurther calcium supplementation.

Over the next 24 hours the dog returned to a normal

activity level. The PCV remained stable at 3538%,

as did the TPP of 5054 g/L (5.05.4 g/dL). The dog

remained in the hospital for a further 3 days and was

discharged on prednisonea (1.8mg/kg, PO, q 12 h),

cyclosporineb (5mg/kg, PO, q 12h), and sucralfatec

(75mg/kg, PO, q 8 h). Subsequent recheck examina-tions were performed through the primary veterinarian

with medications tapered and discontinued over the

following 3 months with no recurrence of the anemia.

Discussion

Therapeutic plasmapheresis has been used for many

years in human medicine as an adjunctive treatment in

a wide variety of conditions. In veterinary medicine the

use of plasmapheresis has been limited by the expense

and availability of technology. Other than 1 case reportdescribing the use of membrane filtration plasma-

pheresis for the treatment of myasthenia gravis,6 previ-

ously documented veterinary case reports have used

a centrifugal technique with a continuous cell separa-

tor.4,5,7 Centrifugal plasma exchange involves the re-

moval of patient blood into an automated system that

continuously separates the plasma and returns the

blood components to the patient. Automated centrifu-gal systems are less user-friendly than newer mem-

brane separation units, and require a more expensive

and less portable machine. The removal of blood cells

from the plasma is also not as complete with this

method. This leads to high numbers of RBC and plate-

lets being lost through discarded plasma, potentially

leading to anemia and thrombocytopenia posttreat-

ment. In recent years technology advancements haveled to membrane separation techniques as an effective

alternative. This technique utilizes a microporous mem-

brane to separate the plasma from the blood, and results

in minimal losses of other blood components. The more

efficient separation of plasma, and replacement of blood

components within the machine, make treatments more

rapid and allow other applications such as double-filtra-

tion plasmapheresis possible. The case reported here is,to the authors knowledge, the first reported veterinary

case describing the use of a continuous membrane sep-

aration technique for treatment of IMHA.

The main indications for plasmapheresis are removal

of autoantibodies, immune complexes, endotoxin, and

toxins.1,11 A single treatment provides a period of de-

creased serum concentration of the target substance

that reduces clinical signs transiently, allowing moretime for medical therapy to take effect.2 The most common

uses for plasmapheresis in human medicine are condi-

tions such as Guillain-Barre syndrome,12,13 Goodpastures

syndrome,13 myasthenia gravis,12,13 autoimmune hemo-

lytic anemias,13 autoimmune thrombocytopenias,12 and

acute hepatic failure as a bridge to liver transplant.13 In

IMHA or immune-mediated thrombocytopenia plasma-

pheresis decreases transfusion requirements, and reduces

Table 1: Quantitative immunoglobulin analysis of serial plasma samples

Pretreatment

(0min)

30

minutes

60

minutes

90

minutes

120

minutes

Posttreatment

(150min)

IgG (g/L) (reference interval5 7.517 g/L) 8 7.8 6 5.2 5.1 5

IgG (mg/dL) (reference interval57501700mg/dL) 800 780 600 520 505 500

IgM (g/L) (reference interval50.451.5 g/L) 1.1 1 0.4 0.3 0.3 0.3

IgM (mg/dL) (reference interval545150mg/dL) 114 100 40 25 25 25

IgG, immunoglobulin G; IgM, immunoglobulin M.

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Plasmapheresis for IMHA in a dog

the risk of life-threatening hemorrhage, respectively.11

In many of these conditions there has been significant

improvement in clinical signs and disease outcome, so

research continues into other potential applications.

Resynthesis of autoantibodies occurs continuously so

immunoglobulin levels will inevitably rebound after

treatment.11 Each plasma exchange reduces plasma IgGconcentration by 3260%.1,6 Achieving the goal IgG

concentration of o4 g/L (o400mg/dL) may requiremultiple treatments.3,14,15 In this case the IgG and IgM

levels were reduced by 37% and 75%, respectively (see

Table 1). The goal as seen in this case is to decrease the

total levels of IgG and IgM to subnormal levels, as they

will rapidly rebound within 2472 hours after treat-

ment. As this is a single case it is difficult to determineif this is significant, although this decrease in immuno-

globulins occurred in conjunction with a slowing of

hemolysis as evidenced by a more stable HCT post-

treatment. An alternative way of characterizing the

response to treatment would be to use flow cytometry

for the detection of IgG on RBC before and after

plasmapheresis. This has been found to be a very sen-

sitive and specific test for IMHA in dogs, and could bea useful adjunctive way to determine the success of

therapy with plasmapheresis.16

Plasmapheresis performed through membrane sepa-

ration as in this case, is achieved in a similar fashion to

hemodialysis. The dialysis catheter placed allows a

high rate of blood flow (minimum of 20mL/min) to be

cycled through an extracorporeal circuit and hemofilter.

The hemofilter separates and removes the plasma con-tinuously before returning the remaining blood cell

components to the patient.17,18 As in hemodialysis, the

filter comprises a semipermeable membrane in a hol-

low fiber design, encased in a polycarbonate casing.19

In plasmapheresis the pore size is much larger than in

hemodialysis allowing molecules up to 13 million Da

to be removed (compared with up to 55,000Da in

hemodialysis).15 Immunoglobulins such as IgG andIgM are 150,000 and 950,000Da, respectively, so they

are effectively cleared by a plasmapheresis filter (but

would not be removed by a hemodialysis filter). Mi-

crofiltration is the process of using both positive and

negative pressure to move fluid across a semipermeable

membrane along a pressure gradient. As the blood

moves through the circuit it passes through the filter at

high pressure before returning to the patient. This fluidmoves across the semipermeable membrane from the

high-pressure blood side, to the low-pressure effluent

side of the circuit. The effluent circuit therefore contains

all the plasma components and molecules that are fil-

tered and will be discarded. A negative pressure is ap-

plied through a pump on the effluent side of the circuit

to help drive this pressure gradient. It is the combina-

tion of the positive pressure on the blood side, and

the negative pressure on the effluent side that creates

the transmembrane pressure (TMP) that drives micro-

filtration. This is a very similar process to ultrafiltration

in hemodialysis, although the larger pore size allows

the passage of larger solutes such as proteins through

the membrane. In hemodialysis the TMP needs to bekept at a higher level to maintain efficient fluid re-

moval. In plasmapheresis fluid removal is not the goal,

so the TMP should be kept o100mm Hg to reduce therisk of hemolysis, which is induced by higher pres-

sures. The other main transport mechanism used in

plasmapheresis is convection, which involves the

movement of solutes through solvent drag with the

water flow that is generated by microfiltration. This al-lows very efficient solute removal without significant

fluid removal. The time required for the plasma

exchange is dependent on the size of the patient, as it

is proportional to the plasma volume removed. In this

patient, we followed the current human recommenda-

tions, replacing twice the patients plasma volume with

fresh frozen plasma. Plasmapheresis in this patient took

2.5 hours, with the plasma delivered to the patient atapproximately 3mL/min. This plasma combines with

the blood that has passed through the filter, to be deliv-

ered back to the patient through the blood return line.

In IMHA the fixation of IgG or IgM antibodies onto

components of the red cell membrane leads to prema-

ture destruction of the RBC.20 The cornerstone of ther-

apy continues to be immunosuppression, although the

response to immunosuppressive therapy is variable. Ifhemolysis is rapid and severe, multiple transfusions

and extended hospitalization may be required.2 In these

cases an adjunctive therapy such as plasmapheresis

may be used to rapidly remove antibodies, immune-

complexes, and activated complement components

until immunosuppression becomes effective.2,13 The

effects are transient as IgG and IgM synthesis will con-

tinue, leading to the need for multiple treatments inmany cases. In this case there was a decrease in both the

IgG and IgM levels posttreatment although repeat sam-

ples were not tested in the following days to evaluate

the rebound phenomenon.

The effectiveness of the plasma exchange relies on

the distribution of IgG and IgM, with 45% of IgG being

distributed intravascularly compared with 70% of

IgM.1 As IgM antibodies predominate in intravascularhemolysis, plasmapheresis may be more effective in

these cases, although large scale clinical trials have not

been performed. In the patient described in this report

there was a greater decrease in the IgM levels consistent

with this increased intravascular distribution.

In human medicine plasmapheresis can be used in

acute hepatic failure patients with hyperbilirubinemia

& Veterinary Emergency and Critical Care Society 2009, doi: 10.1111/j.1476-4431.2009.00431.x378

K.L. Crump & R. Seshadri

to reduce hepatotoxins as a bridge to liver transplantation.

An average decrement of 45% in bilirubin is seen with

each plasmapheresis treatment, with a 100% rebound

within 48 hours.1 This may be useful in patients with

hyperbilirubinemia associated with IMHA, although

double filtration plasmapheresis with immunoadsorbtion,

or techniques such as the molecular adsorbent recircula-tion system used for liver failure, may be more effective at

reducing bilirubin.21 In this patient the preplasmapheresis

bilirubin was mildly elevated, and was normal on

all post-plasmapheresis blood samplings. This was likely

through a combination of both clearance through

the plasmapheresis treatment in addition to decreased

hemolysis.

In humans the most commonly chosen replacementsolutions are human albumin or fresh frozen plasma.

Properties of the replacement solution required include

volume replacement, restoration of oncotic pressure,

and replacement of immunoglobulins and coagulation

factors. In coagulopathic patients fresh frozen plasma is

preferred to replenish coagulation factors. Expense and

risks of disease transmission have meant that human

albumin is often preferred as the primary replacementsolution in humans, although plasma remains the

mainstay of therapy for many disease conditions. Pre-

viously in veterinary medicine fresh frozen plasma has

been utilized, although the use of human albumin for

colloid support has been described.22 Recently concerns

over human albumin causing severe acute and delayed

hypersensitivity reactions have been described in ani-

mals, leading to reservations with its use.23,24 In veter-inary medicine viral transmission through plasma

transfusions has not been described, and anaphylaxis

is uncommonly seen. This made plasma the preferred

replacement solution for this case. There is a risk of

hypovolemia during the initiation of plasmapheresis in

small animals as the total circuit blood volume is 88mL.

As our patient was anemic pre-plasmapheresis and

hypovolemia was a concern, we transfused packed RBCconcurrently.

The choice of anticoagulant is another important

consideration, with the most commonly used anticoag-

ulants being heparin or sodium citrate. Sodium citrate

binds ionized calcium to inhibit the coagulation cascade

in the circuit without affecting the patients coagulation

status.25 Calcium is then delivered to the patient post-

filter through a peripheral catheter to prevent hypo-calcemia. As citrate increases bicarbonate production in

the body, close monitoring for alkalosis is recom-

mended. In this case sodium citrate was successfully

used as the anticoagulant although moderate hypo-

calcemia was created. In this case higher rates of

calcium supplementation should have been considered

to correct the hypocalcemia, although a balance must be

found between anticoagulation and prevention of filter

clotting. This patients calcium levels corrected once the

plasmapheresis was completed and no further calcium

supplementation was required.

Side effects are uncommon in human plasmapheresis

facilities with minimal hemodynamic compromise noted

in their patients. The most common complication is hem-orrhage from the dialysis catheter site that is generally

mild and self-limiting. Vascular access problems can oc-

cur particularly in small patients where placing a large

bore dialysis catheter can be difficult. Inadequate vascu-

lar access can lead to decreased blood flow rates, in-

creased clotting of filters, and interruption in therapy.

Hypersensitivity reactions to the replacement fluid,3

hypotension, citrate-induced hypocalcemia, coagulationabnormalities, and viral transmission from blood prod-

ucts have also been reported.1 We did not note any signs

of hypovolemia or hemodynamic instability during the

plasmapheresis with normal blood pressures, HR, and

pulse quality. The use of plasmapheresis in veterinary

patients requires specialized equipment that is becoming

more available in veterinary referral centers. As experi-

ence grows with the use of this technology, plasma-pheresis may become more accessible and potentially

beneficial in conditions such as IMHA, ITP, and acute

myasthenia gravis crisis. Currently we are limited to us-

ing a filter that makes its use in small patients more

challenging due to the extracorporeal volume. Another

limitation is the cost of treatment, although this may be

comparable to additional days in the ICU with multiple

blood transfusions. The cost of treatment is dependant onthe size of the patient, and the potential need for multiple

sessions of plasmapheresis. In this patient the cost of one

plasmapheresis treatment was approximately $2200USD,

although this cost will vary between different hospitals

and regions. To our knowledge this therapy is not cur-

rently offered in any other facilities, although the filter

can be used with a standard hemodialysis machine. Be-

cause of the lack of controlled trials investigating its effi-cacy it is not currently offered as a standard therapy for

IMHA. It may, however, hold promise as an adjunct to

other therapies if traditional therapy has failed.

This case demonstrates that it is possible to perform

plasmapheresis safely and effectively in the veterinary

ICU. The use of plasmapheresis in this case resulted in

a more stable HCT, and slowed the rate of hemolysis

with a single treatment. Further clinical trials are indi-cated to identify patient populations that may benefit

from this therapy.

Footnotesa Prednisone, West-ward Pharmaceutical Corp, Eatontown, NJ.b Gengraf modified Cyclosporine capsules, USP, Abbott Laboratories,

North Chicago, IL.

& Veterinary Emergency and Critical Care Society 2009, doi: 10.1111/j.1476-4431.2009.00431.x 379

Plasmapheresis for IMHA in a dog

c Sucralfate tablets, Nostrum Laboratories Inc, Kansas City, MO.d Anzemet, dolasetron mesylate injection, Sanofi-Aventis, Bridgewater,

NJ.e Doxycycline Hyclate capsules, West-ward Pharmaceutical Corp.f Pepcid-AC, Johnson & Johnson/Merck, New Brunswick, NJ.g Lovenox, Sanofi Aventis, Quebec, Canada.h Normosol-R, CEVA Laboratories, Overland Park, KA.i HemoCath, MedComp Inc, Harleysville, PA.j Prisma CRRT and TPE Control Unit, Gambro (Renal Care Products) Inc,

Lakewood, CO.k Prisma TPE2000 Set, Gambro (Renal Care Products) Inc; surface area

and volume data per manufacturer specifications.l Anticoagulant acid-citrate-dextrose (ACD) solution, Formula A, Baxter

Healthcare Corporation, Deerfield, IL.m 10% calcium chloride, American Pharmaceutical Partners Inc, Los

Angeles, CA.

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& Veterinary Emergency and Critical Care Society 2009, doi: 10.1111/j.1476-4431.2009.00431.x380

K.L. Crump & R. Seshadri