Answering your questions

Nov. 1, 2008

Total bilirubin critical value

Q What is the critical value for total bilirubin (newborn)
and adult? I am using Roche Integra 800 special bilirubin reagent.

A A critical value is implemented to decrease hospital
error and increase overall patient safety. It is also used to establish the
upper and/or lower limits of life-threatening values as measured in the
clinical lab. Measured analyte results that meet these criteria are
considered urgent and follow a specified lab procedure for notifying the
clinician/caregiver responsible for the patient.

Bilirubin is an analyte that meets the criteria for
reporting critical or threshold values. It is a byproduct of hemoglobin
catabolism and can be measured as total or direct (conjugated to glucuronide)
and reported as total, direct, or indirect. Adult and pediatric critical
values for total serum bilirubin (TSB) have been consistently reported in
the literature to be 15 mg/dL to 20 mg/dL.1,2,3 The variety of
causes for hyperbilirubinemia, includes inherited genetic disorders, biliary
atresia, hemolytic disease, and physiologic jaundice in neonates. The
severity of elevated TSB or progression to hyperbilirubinemia in pediatrics
is of special concern because of its direct role in causing kernicterus if

Newborn jaundice is a common and usually benign condition
that manifests in infants due, in part, to undeveloped enzyme systems for
conjugating bilirubin. Extreme hyperbilirubinemia is rare (1:10,000
newborns); however, if unnoticed, it can result in kernicterus. Kernicterus
is also infrequent but has a significant mortality (~10%) and incurable
long-term morbidity (~70%). The concentration at which this disease can
occur in infants has been reported in the literature to be >20 mg/dL.

Current pediatric monitoring of newborns with jaundice
involves, in part, plotting a newborn's levels over time to track changes in
bilirubin levels for age in hours. An established guideline and nomogram for
designation of risk in healthy newborns at 36 or more weeks has been
established.4 To create this nomogram, serum-bilirubin levels
were obtained from healthy-term newborns before discharge and “risk” zones
were defined to predict the likelihood of a subsequent bilirubin level
exceeding the “high-risk” zone (level associated with kernicterus). For
example, a healthy-term newborn that is 12 hours of age with a serum-bilirubin
level of 10 mg/dL would be considered “high-risk” according to the
established nomogram. This contradicts the “traditional” laboratory criteria
for a critical bilirubin level. Patients with TSB levels in the “high-risk”
zone are evaluated for cause. If necessary, a treatment is introduced and a
repeat of the TSB repeated within the following four to 24 hours.

Determining critical values for a particular analyte are
established from a variety of sources such as the manufacturer's package
insert and from published literature; but, ultimately, it requires the
consensus opinion of physicians and the laboratory medical director.

—Randy Schneider, PhD

Department of Pathology

Medical College of Wisconsin

Milwaukee, WI

—Stanley F. Lo, PhD, DABCC, FACB

Department of Pathology

Medical College of Wisconsin

and Children's Hospital of Wisconsin

Milwaukee, WI


1. Burtis CA, Ashwood ER, Bruns DE, Tietz Textbook of
Clinical Chemistry and Molecular Diagnostics
, 4th ed. WB Saunders:

2. Bhutani VK, Johnson LH, Urgent clinical need for
accurate and precise bilirubin measurements in the United States to
prevent kernicterus. Clin Chem. 2004;50:447-480.

3. Kost GJ, et al. Critical limits for urgent clinical
notification at U.S. Medical Centers. JAMA. 1990;263:704-707.

4. Subcommittee on Hyperbilirubinemia, Management of
Hyperbilirubinemia in the Newborn Infant 35 or more weeks of gestation.

Macrocytosis with normal MCV

Q Is there any reference information to show that moderate
to marked macrocytosis can be present in view of a normal mean red-cell
volume (MCV) of 91 and an RDW (red-blood-cell [RBC] distribution width
[RDW]) of 18.5 with marked targeting. I have seen this set of circumstances
many times over the years; I know that abnormal RBC shapes, such as target
cells, can falsely decrease an MCV.

A MCV indicates the average volume of individual
erythrocytes, which is used to classify RBCs as normocytic, microcytic, and
macrocytic.1 It can be directly measured on automated cell
counters or manually calculated from the hematocrit and RBC count.

If a majority of the cells are larger than normal, then
MCV is increased; if smaller than normal, then MCV is decreased. If there is
significant variation in size with microcytic, normocytic, and macrocytic
cells present, the MCV may be normal because it is the
average of cell size. Therefore, RDW is another helpful parameter to describe
RBC morphology in conjunction with MCV. Increased RDW suggests that RBCs are
heterogeneous in size. Normal MCV with increased RDW means that there are
mixed microcytic and macrocytic RBCs (with or without normocytic RBCs).

To answer the question, I also need to clarify whether
target cells are normocytic, microcytic, or macrocytic. Target cells result
from increased surface-to- volume ratio of the cells. Target cells could be
normocytic, macrocytic, or microcytic, depending on what is the cause of
target-cell formation.2,3,4 For example: 1) normocytic or
macrocytic target cells in liver disease (increased cell membrane lipid
content); 2) microcytic target cells in iron-deficiency anemia and
thalassemia (decreased hemoglobin content in cells); and 3) normocytic
target cells also can be due to artifacts of air drying.

With the above explanation and understanding, these are a
few clinical settings in which you may see normal MCV with moderate to
marked macrocytosis, target cells, and increased RDW: 1) liver disease; 2)
myelodysplastic syndrome; and 3) a combined iron and folate deficiency.

— Guang Fan, MD, PhD

Medical Director, Hematology Service

Department of Pathology

Oregon Health and Science University

Portland, OR


1. MacKenzie SB. Clinical Laboratory Hematology.
Upper Salle River, NJ: Prentice Hall Pearson Education, 2004: 167-187.

2. Glassy EF, ed. Color Atlas of Hematology: An
Illustrated Field Guide Based on Proficiency Testing.
IL: College of American Pathology, 1998:106-109.

3. Lewis SM, Bain BJ, Bates I. Dacie and Lewis
Practical Haematology,
10th ed. Oxford, U.K. Churchill Livingstone;

4. Kaewketthong P, Bunyaratvej A, Barusrux S. Different
cell volume with high target cell population between liver disease and
homogygous hemoglobin E. J Med Assoc Thai. 1992;75 Suppl

UA and culture do not agree

Q What are causes of a urine culture not agreeing with a
urinalysis? The UA shows positive nitrite and positive leukocytes but no
growth on media (plated to blood agar and MacConkey's agar). If the results
do not match, can we take the specimen, which may be 24 hours old or more,
and perform both tests again to check the results?

A Unpreserved urine specimen for culture can be stored for
up to 24 hours at 4oC or, if preserved, the specimen can be
stored at room temperature for up to 24 hours without major effects on
bacterial-culture results.1 If re-testing is performed after 24
hours, results may be questionable as to how they would correlate with the
original specimen culture.

If a patient is already on antibiotics prior to
collecting the urine specimen for analysis, then the urinalysis could be
positive but the culture negative. There are certainly other reasons for
increased white blood cells in the urine that are not related to infection.
Leukocytes in the urine are indicative of injury to the urinary tract, which
may or may not be due to infection. Non-infectious conditions for pyuria
include calculous disease, stricture disease, neoplasm, glomerulonephropathy,
or interstitial cystitis.2 There are also infectious agents that
can cause pyuria which cannot be recovered in the laboratory on blood or
MacConkey's agar plates, such as chlamydia, mycoplasma, and ureaplasma.
Urine nitrites can be falsely elevated due to contamination, air exposure,
or phenazopyridine, and can be falsely negative due to elevated specific
gravity, elevated urobilinogen levels, nitrate reductase-negative bacteria,
pH <6.0, and vitamin C.3

—Susan E. Sharp, PhD (DABMM)

Director of Microbiology

Kaiser Permanente

Pathology Regional Laboratory;

Associate Professor

Oregon Health and Science University

Portland, OR


1. Miller JM. A guide to specimen management in clinical
microbiology. Washington, DC: ASM Press;1996:129.

2. Tierney LM, McPhee SJ, Papadakis MA. Current Medical
Diagnosis and Treatment. 34th ed. Norwalk, CT: Appleton &

3. Simerville JA, Maxted WC, Pahira JJ. Urinalysis: A
Comprehensive Review. Journal of the American Academy of Family
Physicians. March 15, 2005;71(6):1153-62

Best procedure for skin scraping fungi

Q What is the best procedure to use for looking for fungal
elements on slides of nails, skin, and other such samples? Just KOH
(potassium hydroxide), or KOH and a stain? Which stain? One of my techs
worked in a lab that used Chlorazol, which is Chlorazol black E, potassium
hydroxide, and dimethylsulfoxide.

A Two experts provided information about this question.
Glenn D. Roberts, PhD, Division of Clinical Microbiology, Mayo Clinic,
replied, “We have worked with the KOH preparation for over 30 years, using
10% KOH with 10% glycerin added to prevent dehydration. We have no
experience with DMSO (dimethylsulfoxide), and I have never talked to anyone
who uses it. We generally allow the slide to sit for five minutes or more to
hasten clearing of the clinical specimen. The former process was to gently
pass the slide through a gas flame to speed things up, but we no longer do
that. The addition of Calcofluor white to the KOH has markedly improved our
ability to detect organisms in clinical specimens; however, the overall
sensitivity for the method is not as good as we would like.”

Deborah L. Grafelman, president, Delasco, Dermatologic
Lab and Supply, answered, “The purpose of the KOH is to 'clear' (partially
dissolve) the keratin. The fungal structures are resistant to the KOH and
remain; but once the keratin is cleared, the fungal elements become visible
and readily identifiable. KOH in water will do this, but it takes some time
(15 to 20 minutes for clearing). This process can be speeded up by heating;
however, there is a risk of overheating and drying the specimen, or boiling
and splattering the caustic KOH. Additionally, the heating process takes a
few seconds.

“The DMSO, in fact, does carry the KOH into the keratin
much more rapidly; and, unless the specimen is quite thick or is, for
example, a piece of nail plate (either of which may require five to 10
minutes of 'soaking'), the specimen can be examined immediately and is
usually cleared much quicker than even heating KOH and water. The only
disadvantage of which I am aware is the slight garlic odor of DMSO, but only
one user has complained about it in over 20 years of providing the product.
Another characteristic is the fact that these are saturated solutions; and,
if there is any evaporation of the water component, the product may begin to
separate, causing microscopic bubbles on the slide. The 'cure' for this is
to add a few drops of water to the bottle, which reconstitutes the

—Daniel M. Baer, MD

Daniel M. Baer, MD, is professor emeritus of
laboratory medicine at Oregon Health and Science University in Portland,
OR, and a member of
MLO's editorial advisory board.

MLO's “Tips from the Clinical Experts”
provides practical, up-to-date solutions to readers' technical and
clinical issues from a panel of experts in various fields. Readers may
send questions to Dan Baer by e-mail at
[email protected].