Understanding
Protein
Electrophoresis
International Myeloma Foundation
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P
Table of Contents
Introduction
5
WhyIsProteinElectrophoresisImportant? 5
WhatIsMonoclonalProtein?
8
WhatIsProteinElectrophoresis?
11
HowisMonoclonalProteinDetected
andMeasured?
14
HowCanElectrophoresisHelp
withTreatmentDecisions?
16
PracticalRecommendationsforUse
ofSPEPandUPEP
17
WillInsuranceCovertheCost
ofProteinElectrophoresis?
19
AdditionalQuestions
20
AbouttheIMF
21
Glossary
24
Sponsored by an unrestricted educational grant from Sebia.
©2011, International Myeloma Foundation, North Hol ywood, California (vii11)
Introduction
You received this booklet to learn more
about a type of laboratory test cal ed Protein
Electrophoresis (PEP). After reading this
booklet, you should be able to answer the
fol owing questions:
n What is protein electrophoresis?
n Why are the values of this test important
for someone with multiple myeloma?
n How does this test help with diagnosis
and monitoring response to treatment of
multiple myeloma?
This booklet is intended to provide you with
general information only. It is not meant to
replace the advice of your doctor or nurse
who can answer questions related to your
specific treatment plan. The definitions of al
words in bold print are found in the glossary
at the end of the booklet.
Why is Protein Electrophoresis
Important?
Why have a booklet about protein electro-
phoresis? The main reason is that production
of a single, monoclonal protein (M-protein) is
a characteristic feature of multiple myeloma.
The test used to measure the amount of
monoclonal protein in the blood or urine is
protein electrophoresis.
This M-protein is manufactured (synthesized)
by malignant plasma cel s (or myeloma
5
cel s). The amount of protein produced and
Measurement of M-protein
released into the serum (the liquid part of
with Electrophoresis
the blood that is left after the blood cel s
If the M-protein, which is characteristic of
are removed), and sometimes into the urine,
myeloma, is present in serum, the electro-
reflects the amount of myeloma present in the
phoresis test for it is cal ed Serum Protein
body at any given time. This protein, which
ElectroPhoresis (SPEP). Likewise, when mono-
is released into the serum or urine, is cal ed
clonal protein is found in urine, the test for it
a serum or urine tumor marker.
is cal ed Urine Protein ElectroPhoresis (UPEP).
Only a very few cancers have this type
To measure the amount of the monoclonal
of a marker which, in this case, makes it
or M-protein one needs two pieces of
possible to assess the amount of myeloma
information:
at the time of initial diagnosis and track the
n What is the total amount of protein
amount of myeloma throughout the course
in the serum or urine?
of the disease. One can look at response
to treatment, depth of remission, and, if
n What percentage of the total is the
necessary, the patient's relapse using exact
M-protein?
numbers, which is a unique advantage. For
The crucial information comes from SPEP
example, we can determine if a response
and/or UPEP. By calculating the size of the
is: Partial (PR)=50% reduction of M-protein;
"spike", which depicts the amount of mono-
very good partial (VGPR)=90% reduction
clonal protein (done by measuring the area
of M-protein; or complete (CR)=no protein
between the top of the spike and the baseline
detected. We can also identify a 25%
of the graph), one gets the percentage of the
increase in protein level, which we cal
total protein that represents the M-protein.
relapse.
For example:
n Spike is 60% of total protein
n Total protein=12 grams/deciliter (g/dL)
n Spike level=7.2 g/dL (60% of 12)
Both the total protein and percentage of
monoclonal protein change over time.
With response to treatment, the spike can
drop to 40%, and total protein to 9 g/dL,
for example. This gives a spike level of
Figure 1. Representation of a normal
3.6 g/dL, which is a 50% drop in M-protein,
Serum Protein Electrophoresis result
or a partial response.
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of an immunoglobulin molecule. Normal
plasma cel s produce immunoglobulins, which
are the antibodies necessary to fight infec-
tion. The abnormal plasma cel s "myeloma
cel s" present in myeloma patients do not
produce antibodies in response to infection.
Instead, they produce a monoclonal immu-
noglobulin molecule that cannot function as
an antibody. This immunoglobulin can be
comprised of: two heavy chains and two
light chains (see Figure 2); light chains only;
or fragments/combinations of this immuno-
globulin molecule that are unique to each
particular myeloma patient.
Immunoglobulins can be formed from one of
This type of serial measurement and assess-
five possible types of heavy chains (referred
ment is key to al myeloma disease moni-
to as IgG, IgA, IgM, IgD and IgE) and
toring. This is why SPEP and UPEP are so
two types of light chains (kappa, , and
important.
lambda, ). Therefore, there are ten possible
An additional test, cal ed immunofixation, is
combinations of heavy and light chains:
used to determine the type of M-protein. This
IgG kappa, IgG lambda, IgA kappa, IgA
is important at the time of baseline diagnosis
lambda, IgM kappa, and so on. Protein
and at the point of maximum response with
electrophoresis is able to detect al of them.
treatment. If there is a complete response
(CR), the immunofixation test is completely
negative*; that is, no monoclonal protein
can be identified.
What Is Monoclonal Protein?
Monoclonal proteins are immunoglobulin
(abbreviated as "Ig") molecules or parts
* How response is defined has been addressed by the IMF's International
Myeloma Working Group (IMWG) and their publication of Uniform
Response Criteria. Please refer to the IMF publication Concise Review of
the Disease and Treatment Options, Table 7, for further information.
Figure 2. Immunoglobulin structure
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Plasma cel s produce heavy chains and light
What Is Protein Electrophoresis?
chains separately, and these chains are later
assembled to form an intact immunoglobulin.
Serum Protein Electrophoresis
For some reason, plasma cel s usual y pro-
Serum contains a variety of dif erent proteins
duce a higher number of light chains than of
that wil be separated by electrophoresis into
heavy chains. Thus, once the intact immuno-
five or six fractions (according to the method
globulins are formed, there stil wil be some
used by the laboratory). These fractions
light chains remaining. These light chains are
(also referred to as "zones" or "regions")
cal ed "free light chains" (the light chains that
are cal ed Albumin, Alpha 1, Alpha 2, Beta
participate in the formation of intact immu-
(which can be separated into Beta 1 and
noglobulins are cal ed "bound light chains,"
Beta 2), and Gamma. Polyclonal (normal)
because they are bound to heavy chains).
immunoglobulins are found mostly in the
The excess of free light chains enters the
Gamma zone. Hence, they are known as
bloodstream and is subsequently filtered by
gamma globulins.
the kidneys. The kidneys are then able to
The diverse, normal immunoglobulins pres-
resorb these free light chains and recycle the
ent in serum dif er slightly from each other
amino acids (building blocks that make up
in their structure and electrical charge. For
al proteins in the body). But when a mono-
that reason, when they are subjected to elec-
clonal protein is present in the serum, the
trophoresis, they form a large zone, which
amount of monoclonal free light chains can
is spread out and symmetrical, without any
become too high for the kidneys to resorb. In
visible deformation.
this case, monoclonal free light chains are fil-
Monoclonal proteins are produced by one
tered through the kidneys, and those that are
clone of plasma cel s, and thus al the mole-
not resorbed can be found in the urine; they
cules are identical and have the same electri-
are then cal ed Bence Jones protein (named
cal charge. That is why on electrophoresis a
after the physician who first described them).
monoclonal protein wil migrate as a narrow
spike (this spike appears most often in the
gamma zone, but sometimes can be present
in Beta 2 or Beta 1, or even the Alpha 2
zone, although the lat er is very rare).
Serum electrophoresis can be used to search
for a monoclonal protein as wel as to moni-
tor the amount of monoclonal protein.
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monoclonal protein may be suspected. It is
necessary then to confirm its presence and to
determine its type by identifying which types
of heavy chains and light chains are involved
in its structure. Knowing the type of M-protein
is important in establishing a diagnosis and
in monitoring the patient.
Figure 2. Representation of an abnormal
To do this, another method of electropho-
Serum Protein Electrophoresis result,
resis, cal ed immunofixation (or IFE, for
with myeloma cells producing the M-protein,
ImmunoFixation Electrophoresis), wil be
creating an M-spike in the gamma zone.
used. Immunofixation methods are more
Urine Protein Electrophoresis
sensitive to the presence of faint monoclo-
nal proteins and may detect them even if
The kidney acts as a filter, eliminating only
electrophoresis does not show any visible
a few molecules and leaving most of the
proteins in the bloodstream. Although some
smal proteins do pass through the kidney fil-
ter, they are later resorbed and recycled into
amino acids. Thus, normal y urine contains
only traces of proteins.
When monoclonal protein is present in
serum, often the excess of free light chains
wil be found in the urine as Bence Jones
protein (it wil look like a narrow spike, usu-
al y in the Gamma or Beta zone).
Urine electrophoresis is used to search for
Bence Jones protein and to monitor its con-
centration. It can also help to assess kidney
damage (which is a common complication of
multiple myeloma).
Serum and Urine Immunofixation
Once a narrow spike of protein is detected
by protein electrophoresis, the presence of
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abnormality. But immunofixation does not
visual y. A monoclonal protein is suspected
determine how much M-protein is present.
when the electrophoresis pat ern exhibits an
Therefore, both methods are used together:
additional spike, or when a normal fraction
electrophoresis to detect the monoclonal pro-
presents an abnormal shape or is unusual y
tein and to quantify it, and immunofixation to
increased. These abnormalities are usual y
identify its type.
detected in the Gamma or Beta regions.
To confirm the presence of the monoclonal
How is Monoclonal Protein
protein and to identify its heavy and light
Detected and Measured?
chains, the serum is further analyzed by
immunofixation. Once the presence of a
As with other tests, the values calculated by
monoclonal protein is confirmed, its amount
electrophoresis are then compared to the
wil be quantified using the electrophoresis
laboratory's reference values (values that
curve. To do this, the laboratory wil mark
a laboratory has determined by analyz-
the beginning and the end of the monoclo-
ing samples coming from healthy people).
nal peak on the curve, and calculate what
Electrophoresis fractions are also analyzed
percentage of the total area under the curve
the peak occupies. Using this value and the
results of another biochemical test, total pro-
tein concentration, the amount of monoclonal
protein in g/dL can be calculated. (See the
example on p. 7.)
This value wil be used by your doctor, in
conjunction with other data, to establish a
diagnosis. Later, it wil be compared with
the values obtained by subsequent electro-
phoresis results to monitor the evolution of
the disease and the response to treatment,
as recommended by International Myeloma
Working Group guidelines.
According to these guidelines, when a
monoclonal protein can be detected and
measured, by electrophoresis, monitoring
should be performed using serum and/or
urine electrophoresis. When the monoclonal
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protein is undetectable by electrophoresis or
important in terms of response to treatment
is too smal to be measured your doctor wil
as wel as potential progression or relapse.
use another test, the free light chain assay, to
The ways in which SPEP and UPEP help are:
monitor your condition.*
1. Baseline assessment of the amount and
type of the myeloma.
How Can Protein Electrophoresis
2. Serial monitoring to document the speed
Help with Treatment Decisions?
and level of response.
As previously mentioned, the ability to pre-
3. Assessment of possible disease progres-
cisely measure the amount of monoclonal
sion or relapse.
protein by SPEP and/or UPEP and to know
Note: Progressive Disease (PD) = 25%
the exact level of the myeloma tumor burden
increase** (from low point). PD can also
is a huge advantage. Disease monitoring in
be confirmed by occurrence of any one of
the absence of a spike on SPEP or UPEP can
the "CRAB" features***.
be quite dif icult.
The level of M-component on SPEP and
Practical Recommendations
UPEP reflects the amount of myeloma pres-
for use of SPEP and UPEP
ent. However, it is very important to realize
When new myeloma treatment is started, the
that each patient is dif erent. At the time of
M-protein level in blood (serum) and/or urine
diagnosis, some patients have, for example,
should be measured each month (or each
a very high spike in the serum but not such
cycle of treatment, normal y at 36 week
a high level of myeloma in the bone marrow
intervals) using SPEP and/or UPEP. Regular
or bones. And the opposite is also true: some
testing is required to assess the impact of
patients can have a low spike, but a lot of
treatment. If the treatment is not working
myeloma cel s. Thus, at diagnosis, it is very
wel , this wil be evident within 23 months:
important to correlate the spike level with the
the SPEP/UPEP M-protein levels would drop
amount of myeloma in an individual patient.
by <25% or increase by 25% (progressive
If the spike is low, this can be especial y
disease [PD]). At this point a change in treat-
important, since smal changes can be more
ment may be required. This is an important
* For further guidance on disease monitoring, refer to the fol owing IMWG
publications. Al are available at myeloma.org. IMWG Uniform Response
**Two consecutive readings are required to confirm progression or relapse.
Criteria, IMWG Guidelines for Serum Free Light Chain Analysis (update in
progress), and IMWG Guidelines for the Diagnostic Work-up of Myeloma.
***See other IMF publications and the IMF web site at myeloma.org, or
Additional y, refer to the 2011/2012 edition of the IMF publication Concise
cal the IMF hotline at 800-452-2873 (in the U.S. and Canada) for an
Review of the Disease and Treatment Options.
explanation of "CRAB" features and additional details.
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time to discuss the results and treatment
computer program, which enables one to
options with the doctor.
create tables, graphs, and flow charts of test
If the treatment is working wel , frequently
results over time. This facilitates patient-doctor
there wil have been substantial improve-
discussions and planning.
ment (reduction) in the M-protein levels on
SPEP/UPEP within the first 23 months.
Will Insurance Cover the Cost
As noted above, the level of response can
of Protein Electrophoresis?
be quantified as 50% (partial response
Protein electrophoresis is a standard,
[PR]); 90% (very good partial response
accepted test which is normal y covered by
[VGPR]); 100% (complete response [CR]).
Medicare and most insurance programs.
The classification of CR also requires that no
M-protein is detectible using immunofixation
However, like al testing, it may be that insur-
(IFE) and that a bone marrow test shows no
ance covers only part (for example, 80%) of
evidence of myeloma. Achieving the higher
the cost. It is important to be aware of what
levels of response such as VGPR and CR wil
patient cost is incurred with each test.
typical y take at least another 23 months of
treatment (total of 46 months at that point).
Several additional months of therapy may be
required to achieve maximum response.
Further decisions are required with respect
to additional treatment options, such as an
autologous stem cel transplant (ASCT) or
consolidation therapy to maximal y improve
the response. Thereafter, there can be a deci-
sion to begin maintenance therapy, and this
is best discussed with your physician.
Once a stable level of response is reached,
the frequency of monitoring can be reduced
to every 23 months, which can continue
on an ongoing basis unless new issues
emerge. It is most helpful if direct discussions
with your doctor occur about the SPEP/
UPEP results as they become available. The
IMF of ers the free Myeloma ManagerTM
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If cost is an issue, discuss the urgency and/or
About the IMF
needed frequency of testing with your treat-
ing physician. If the myeloma is stable, less
"One person can make a dif erence,
frequent testing may be acceptable.
Two can make a miracle."
Additional Questions
Brian D. Novis
IMF Founder
We suggest the discussion of the fol owing
Myeloma is a lit le-known, complex, and
questions with your doctor to provide a
often misdiagnosed bone marrow cancer that
bet er understanding of your electrophoresis
at acks and destroys bone. Myeloma af ects
results:
approximately 75,000 to 100,000 people
n What are the normal ranges for
in the United States, with approximately
immunoglobulin levels on SPEP at this lab,
20,000 new cases diagnosed each year.
and how do my results compare?
Although there is presently no known cure for
n How much variation in monthly SPEP and/or
myeloma, doctors have many approaches to
UPEP results is considered within normal
help myeloma patients live bet er and longer.
limits? At what point is an increase or
decrease considered significant?
The International Myeloma Foundation (IMF)
n If there is no detectable monoclonal protein
was founded in 1990 by Brian and Susie
by electrophoresis, does that always mean
Novis shortly after Brian's myeloma diagno-
I am in complete response?
sis at the age of 33. It was Brian's dream that
future patients would have easy access to
n Is it necessary to do both serum and urine
electrophoresis?
medical information and emotional support
throughout their bat le with myeloma. He
n Should patients with MGUS be monitored
established the IMF with the 3 goals of treat-
with serum and urine electrophoresis? If so,
how often?
ment, education, and research. He sought
to provide a broad spectrum of services for
patients, their families, friends, and health
care providers. Although Brian died 4 years
after his initial diagnosis, his dream didn't.
Today, the IMF reaches out to an interna-
tional membership of more than 195,000.
The IMF was the first organization dedicated
solely to myeloma, and today it remains
the largest.
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The IMF provides programs and services to
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(for example, a G type) combined with two light
Glossary
chains, which are either kappa or lambda. Therefore,
Antibody: A protein produced by certain white blood
the two most common subtypes of myeloma have
cel s (plasma cel s) to fight infection and disease in
identical heavy chains (i.e. IgG kappa and IgG
the form of antigens such as bacteria, viruses, toxins,
lambda). The terms heavy and light refer to the size or
or tumors. Each antibody can bind only to a specific
molecular weight of the protein, with the heavy chains
antigen. The purpose of this binding is to help destroy
being larger than the light chains.
the antigen.
Since the light chains are smal er, they are more likely
Antibodies can work in several ways, depending on
to leak out into the urine, resulting in urine Bence
the nature of the antigen. Some antibodies disable
Jones protein.
antigens directly. Others make the antigen more
IgD, IgE: Two types of myeloma that occur less
vulnerable to destruction by other white blood cel s.
frequently.
Bence Jones protein: A myeloma monoclonal protein
IgM: Usual y associated with Waldenstrom's macro-
present in urine. The amount of Bence Jones protein is
globulemia. In rare cases it can be a type of myeloma.
expressed in terms of grams per 24 hours. Normal y
a very smal amount of protein (<0.1 g/24 h) can be
Immunofixation: An immunologic test of the serum
present in the urine, but this is albumin rather than
or urine used to identify proteins in the blood. For
Bence Jones protein.
myeloma patients, it enables the doctor to identify the
M-protein type (usual y IgG, IgA, kappa, or lambda).
The presence of any Bence Jones protein is abnormal.
The most sensitive routine immunostaining technique,
CRAB: C = elevated Calcium, R= Renal (kidney)
it identifies the exact heavy and light chain type
dysfunction, A = Anemia, B = Bone disease
of M-protein.
Electrophoresis: A laboratory test in which a patient's
Immunoglobulin (Ig): A protein produced by plasma
serum (blood) or urine molecules are subjected to
cel s; an essential part of the body's immune system.
separation according to their size and electrical
Immunoglobulins at ach to foreign substances which
charge. For myeloma patients, electrophoresis of
have entered the body (antigens such as bacteria,
the blood or urine al ows both the calculation of the
viruses, fungi) and assist in destroying them. The
amount of myeloma protein (M-protein) as wel as the
classes of immunoglobulins are IgA, IgG, IgM, IgD,
identification of the specific M-spike characteristic for
and IgE.
each patient. Electrophoresis is used as a tool both for
Monoclonal: A clone or duplicate of a single cel .
diagnosis and for monitoring.
Myeloma develops from a single malignant plasma
Free light chains: A portion of the monoclonal protein
cel (monoclone). The type of myeloma protein pro-
of light molecular weight that can be measured in a
duced is also monoclonal; a single form rather than
sensitive assay, the Freelite® test.
many forms (polyclonal). The important practical
aspect of a monoclonal protein is that it shows up as a
IgG, IgA: The two most common types of myeloma.
sharp spike (M spike) in the serum electrophoresis test.
The G and the A refer to the type of protein produced
by the myeloma cel s. The myeloma protein, which
is an immunoglobulin, consists of two heavy chains,
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M-protein (M-spike): Antibodies or parts of antibod-
ies found in unusual y large amounts in the blood
or urine of multiple myeloma patients. M-spike refers
to the sharp pat ern that occurs on protein electro-
phoresis when an M-protein is present. Synonymous
with monoclonal protein and myeloma protein. (See
monoclonal above.)
Plasma cells: Special white blood cel s that produce
antibodies. The plasma cel is the malignant cel in
myeloma. Normal plasma cel s produce antibodies
to fight infection. In myeloma, malignant plasma cel s
produce large amounts of abnormal antibodies that
lack the capability to fight infection.
The abnormal antibodies are the monoclonal pro-
tein, or M-protein. Plasma cel s also produce other
chemicals that can cause organ and tissue damage
(i.e., anemia, kidney damage, and nerve damage).
Reagent: a chemical substance known to react in a
specific way. A reagent is used to detect or synthesize
another substance in a chemical reaction.
Relapse: The reappearance of signs and symptoms of
a disease after a period of improvement.
Remission or response: Complete or partial disappear-
ance of the signs and symptoms of cancer. Remission
and response are used interchangeably.
Tumor marker: A substance in blood or other body
fluids that may suggest that a person has cancer. In
the case of myeloma, M-protein is a tumor marker; it
is an indirect way to gauge the number and activity
of myeloma cel s.
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