Answering your questions

Dec. 1, 2009

Q Most of our
testing is closed-system sampling, but there are a few things for which
we have to take off the caps — for example, putting the uncapped tube on
the DXI, then recapping it for the LX. I know we spin these down, and
the blood goes to the bottom; but is there a possibility of
contamination when not using the original cap? Some of the techs save
the caps, and some recap, using any cap on any tube. They are also
stored this way. If we had to use the specimen for “blood in lab,” might
these samples be contaminated?

A You make a valid point
of potential contamination. Although there is nothing in the literature
that proves or disproves your theory, mixing stoppers with other
patients' tubes indiscriminately is taking a cavalier approach to
maintaining specimen integrity. Whenever there is a chance of
introducing a variable of unknown influence, it should be avoided. A
better approach would be to seal tubes for storage with paraffin rather
than the stopper from another patient's sample. If the sample needs to
be recapped so it can be put onto a cap-piercing instrument, the
original cap or a new cap from an unused tube should be used.

—Dennis J.
Ernst, MT(ASCP)


Center for Phlebotomy Education

Corydon, IL

Q Can an
Rh-positive parent have an Rh-negative child?

A I performed blood
grouping on the 2-month-old child of a patient who is Rhesus positive.
The child turned up to be Rhesus negative. Is it possible that a parent
who is Rhesus positive can have a Rhesus negative child? Or could this
be a technical error?

The Rhesus (Rh) blood group system is a highly
polymorphic blood system with over 40 antigens, including the D antigen.
It is the second most important blood group system after the ABO group
system. This is due to the fact that the D antigen is the most
immunogenic red blood cell (RBC) antigen (after the A and B antigens of
the ABO group system) and is involved in hemolytic disease of the fetus
and newborn as well as extravascular transfusion reactions.

The major antigens of the Rh system are encoded by
RHD and RHCE genes on chromosome 1. The RHD gene codes for the D antigen,
while the RHCE gene codes are for the C, E, c, and e antigens in
various combinations (ce, cE, Ce, or CE).1 The frequency of the
five major Rh antigens in various ethnic groups is listed below:2

Rh antigen


D 85% 92% 99%
C 68% 27% 93%
E 29% 22% 39%
c 80% 96% 47%
e 98% 98% 96%

The terms “Rh-positive” and “Rh-negative” refer only
to the presence or absence of the D antigen on the RBC. In the Caucasian
population, this is due to a deletion of the RHD gene.1 In black
and Asian populations, this phenotype is most often due to various other
mechanisms such as premature stop codons or hybrid genes.3

An Rh-positive parent can have an Rh-negative child.
Individuals inherit two copies of the RHD gene, one from each parent. It is
expressed in a dominant manner. If the parents are both homozygous for RHD,
then the child will be Rh-positive. If both parents are heterozygous for
RHD, they would both type as Rh-positive. Roughly 75% of their children,
however, would be Rh-positive (homozygous or heterozygous for RHD); and 25%
of their offspring would be Rh-negative (homozygous for the RHD deletion).
Interestingly, there are racial variations in the expression of D.
Therefore, this situation would be more common in a Caucasian or black
population than in an Asian population because of the frequency of the
wildtype RHD in the respective populations.

Although less likely, other possibilities such as a
decreased expression of D (weak D phenotype) or expression of portions of
the D antigen (partial D phenotype) can cause false-negative D typing and
could also be an explanation. Unlike weak D individuals, partial D
individuals can make anti-D if transfused and alloimmunized with Rh-positive
blood or exposed during pregnancy.1 These possibilities could be
proven by additional testing of the child's red cells using 37^0C incubation
and an anti-human globulin reagent.1

Finally, incorrect determination of Rh-D phenotype of
an individual can result from technical problems in the laboratory. Failure
to add the anti-D reagent, heavy cell suspension, presence of contaminants
in the sample, or under-centrifugation/incorrect calibration during testing
can result in false-negative reactions. Sample mix-ups and/or clerical
errors also can lead to incorrect determination or interpretation of

—Karafa S.W. Badjie, MS,

Supervisor of Transfusion Laboratory

Mayo Clinic College of Medicine

—Eapen K. Jacob, MD

Division of Transfusion Medicine

Department of

Laboratory Medicine and Pathology

Mayo Clinic College of Medicine

Rochester, MN


  1. Roback JD, Combs MR, Grossman BJ, Hillyer CD. Technical Manual.
    16th ed.
    Bethesda, MD: AABB; 2008.
  2. Dean L. Blood Groups and Red Cell Antigens. Bethesda, MD:
    National Center for Biotechnology; Information. U.S. National Library of
    Accessed September 4, 2009.
  3. Westhoff CM. The Rh blood group system in review: A new face for the
    next decade. Transfusion. 2004; 44:1663-1673.

Color, clarity of urine is important

Q My clinic is part
of a four-clinic group that shares an electronic medical records (EMR)
database. One of the clinics does not want to report color and clarity for
urinalysis (UA). Are these required elements of a UA? Can you give me any
evidence that supports reporting these visual observations?

A Your question regarding
whether labs are
required to report color and clarity as part of a urinalysis is an
intriguing one. As far as we could find, there are no CLIA regulations which
require the inclusion of color and turbidity in a urinalysis report. The
College of American Pathologists (CAP) inspection checklists for labs
performing urinalysis do not list any requirement for inclusion of color or
turbidity in reporting out urinalysis results. The guidelines for urinalysis
published by the Clinical and Laboratory Standards Institute, however,
include the following statement regarding the reporting of color and clarity
of urine: “In consultation with clinicians, each laboratory should determine
whether or not these physical parameters should be part of the routine
urinalysis. However, any unusual color, clarity, or odor should be noted on
the report.”1

The guidelines set forth by the European
Confederation of Laboratory Medicine in 2000 recommend the following:
“Visual inspection is often undertaken and reported by the patient but
should not be omitted by the laboratory.”2 Color and
clarity/turbidity are almost always reported by urinalysis labs for a very
important reason: They can provide valuable additional information regarding
the pathophysiologic state of the patient.

Though rarely sensitive or specific, the color and
clarity of urine provides a quick, cheap, and easy method of assessing
health. In fact, the visual examination of human urine for the purposes of
diagnosing disease dates back as far as 300 BC, when Hippocrates first
associated the appearance of bubbles on urine with renal disease.3
Routine visual inspection of urine, known as uroscopy or “water casting,”
was conducted by physicians in medieval Europe, and the urine flask became
the emblem of medieval medicine. The first guidelines for the use of urine
as a diagnostic aid were established around 900 AD by a Jewish physician by
the name of Isaac Judaeus. Under the Jerusalem Code of 1090, failure by a
physician to examine a patient's urine resulted in his punishment by public
beatings. The practice of uroscopy continued on well into the 17th century,
when it was used routinely to diagnose a condition known as chlorosis or
“love sickness,” as well as chastity in young women. Urinalysis has advanced
somewhat since then, yet the visual appearance of urine continues to provide
information which may not be provided by other routine urinalysis tests.

The factors and conditions leading to abnormal urine
appearance are numerous and complex, and references are available which
cover this topic in extensive detail.2,4 Urine from a healthy
individual should normally appear in a range between pale yellow to amber.
The yellow color is due to the presence of urochrome, a product of
endogenous metabolism. The amount excreted can vary depending on the
person's metabolic state, with increased urochrome production during fasting
states. The darkness of one's urine also varies markedly, depending upon the
hydration state of the individual with dilute urines being nearly clear and
concentrated ones very dark.

Disease states can also change the appearance of
urine. For example, a dark yellow to orange color may signify the presence
of bilirubin, which is indicative of liver disease and/or biliary-tract
obstruction, while brown and/or black urine may be caused by the presence of
melanin or homogentisic acid. Importantly, neither melanin nor homogentisic
acid are detected in a routine urinalysis, therefore careful attention to
the color of urine can provide an important clue to an underlying disease
process. Other examples include blue or green urine, which can indicate the
presence of a urinary-tract infection by Pseudomonas and is caused by
production of indican. Red urine may be caused by the presence of red blood
cells, hemoglobin, or myoglobin. Clear red urine in the absence of red blood
cells may be indicative of in vivo hemolysis or rhabdomyolysis.

Abnormal urine color may also be a result of diet or
ingestion of medications, such as red urine resulting from red beets, and
brown/black urine due to levodopa or phenol derivatives. Some pathologic
causes of turbidity in urine include the presence of red blood cells,
leukocytes, bacteria, yeast, epithelial cells, crystals, lymph, and lipids.
Non-pathologic causes may include the presence of squamous epithelial cells,
mucus, semen, fecal contamination, talcum powder, vaginal creams, or urinary
crystals. It is important to note that color and turbidity are neither
sensitive nor specific enough to be used solely as a method for diagnosing
disease, and should always be interpreted in conjunction with other
laboratory results and clinical presentation.

Another factor to keep in mind is that an abnormal
urine appearance is often what leads a patient to seek medical attention.
Thus, it is important to have the urine sample observed by trained personnel
under a good light source against a white background and documented in the
patient's laboratory results. Before a decision is made to end reporting of
these parameters, we strongly recommend that the laboratory/medical director
have a discussion with the clinicians ordering the urinalysis and inquire as
to the reasons why they do not want color and/or clarity reported. In our
opinion, they should always be reported.

—Kazunori Murata, PhD

Fellow, Clinical Chemistry

—John Lieske, MD


Renal Function Laboratory

Department of

Laboratory Medicine and Pathology
Mayo Clinic
Rochester, MN


  1. CLSI. Urinalysis; Approved Guideline-Third Edition. CLSI Document
    GP16-A3. Wayne, PA. Clinical and Laboratory Standards Institute; 2009.
  2. European urinalysis guidelines. Scand J Clin Lab Invest Suppl.
  3. Berger D. A brief history of medical diagnosis and the birth of the
    clinical laboratory. Part 1-Ancient times through the 19th century.
  4. Strasinger SK, DiLorenzo MS.
    Urinalysis and body fluids. 5th ed. Philadelphia, PA: FA Davis;

Brad S. Karon, MD, PhD, is assistant professor of
laboratory medicine and pathology, and director of the Hospital Clinical
Laboratories, point-of-care testing, and phlebotomy services at Mayo
Clinic in Rochester, MN.