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Laboratory Perspectives in the Diagnosis and Monitoring in Plasma Cell/B Lymphoid Dyscrasias

Jerry Katzmann, Ph.D.

Dept. Laboratory Medicine & Pathology

Katzmann, Ph.D. Dept. Laboratory Medicine & Pathology Mayo Clinic June 8, 2012, Royal Free London Department

Mayo Clinic

June 8, 2012, Royal Free London
June 8, 2012, Royal Free London
& Pathology Mayo Clinic June 8, 2012, Royal Free London Department of Laboratory Medicine and Pathology
& Pathology Mayo Clinic June 8, 2012, Royal Free London Department of Laboratory Medicine and Pathology

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

DISCLOSURES:

Travel expenses, The Binding Site, Ltd. TBS FLC & HLC immunoassay reagents at no cost for clinical studies

HLC immunoassay reagents at no cost for clinical studies Department of Laboratory Medicine and Pathology ©
HLC immunoassay reagents at no cost for clinical studies Department of Laboratory Medicine and Pathology ©
HLC immunoassay reagents at no cost for clinical studies Department of Laboratory Medicine and Pathology ©
HLC immunoassay reagents at no cost for clinical studies Department of Laboratory Medicine and Pathology ©
HLC immunoassay reagents at no cost for clinical studies Department of Laboratory Medicine and Pathology ©

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for Medical Education and Research.
Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for Medical Education and Research.
Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for Medical Education and Research.

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Serum Protein Electrophoresis & Immunofixation Electrophoresis

Normal Serum

& Immunofixation Electrophoresis Normal Serum M M S e r u m Department of Laboratory Medicine

MM Serum

Electrophoresis Normal Serum M M S e r u m Department of Laboratory Medicine and Pathology

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Monoclonal Gammopathies: Mayo Clinic 2010 (n=1,609)

Monoclonal Gammopathies: Mayo Clinic 2010 (n=1,609) Lymphoproliferative 2% (35) SMM 4% (66) Amyloidosis (AL) 9% (152)

Lymphoproliferative 2% (35) SMM 4% (66)

Amyloidosis (AL) 9% (152)

Multiple myeloma 19% (307) MGUS 52% (836)
Multiple
myeloma
19% (307)
MGUS
52% (836)

Solitary or extra- medullary 2% (33)

Macro 4% (56)

Other 8% (124)

or extra- medullary 2% (33) Macro 4% (56) Other 8% (124) Department of Laboratory Medicine and

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Laboratory problems for electrophoretic diagnosis and monitoring of plasma cell dyscrasias

Free light chains: identification & monitoring

FLC quantitation

Monitoring very large M-spikes on agarose

Ig and/or HLC quantitation

Monitoring small M-proteins in γ or β fractions

Monitoring small M-proteins in γ or β fractions • HLC quantitation Broadly migrating M-proteins :

HLC quantitation

Broadly migrating M-proteins : identification & monitoring

HLC quantitation

: identification & monitoring • HLC quantitation Department of Laboratory Medicine and Pathology © 2010 Mayo

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Serum & Urine Protein Electrophoresis and Immunofixation Electrophoresis

LCMM Serum

and Immunofixation Electrophoresis LCMM Serum LCMM Urine Department of Laboratory Medicine and Pathology
and Immunofixation Electrophoresis LCMM Serum LCMM Urine Department of Laboratory Medicine and Pathology
and Immunofixation Electrophoresis LCMM Serum LCMM Urine Department of Laboratory Medicine and Pathology

LCMM Urine

and Immunofixation Electrophoresis LCMM Serum LCMM Urine Department of Laboratory Medicine and Pathology © 2010 Mayo

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

1027 Newly Diagnosed Multiple Myeloma:

Monoclonal Serum Proteins

IgG

52%

IgA

21%

IgM

0.5%

IgD

2%

Biclonal

2%

Free Light Chain only Nonsecretory Myeloma

Free Light Chain only Nonsecretory Myeloma

20%

2.8%

Free Light Chain only Nonsecretory Myeloma 20% 2.8% Kyle et. al., Mayo Clin Proc 2003 Department
Free Light Chain only Nonsecretory Myeloma 20% 2.8% Kyle et. al., Mayo Clin Proc 2003 Department

Kyle et. al., Mayo Clin Proc 2003

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Serum & Urine Protein Electrophoresis and Immunofixation Electrophoresis:

Primary Amyloid

and Immunofixation Electrophoresis: Primary Amyloid AL, Serum AL, Urine Department of Laboratory Medicine and
and Immunofixation Electrophoresis: Primary Amyloid AL, Serum AL, Urine Department of Laboratory Medicine and
and Immunofixation Electrophoresis: Primary Amyloid AL, Serum AL, Urine Department of Laboratory Medicine and

AL, Serum

Immunofixation Electrophoresis: Primary Amyloid AL, Serum AL, Urine Department of Laboratory Medicine and Pathology ©

AL, Urine

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Principle of FLC assay

Principle of FLC assay exposed surface antigen binding sites exposed surface hidden surface hinge region

exposed surface

Principle of FLC assay exposed surface antigen binding sites exposed surface hidden surface hinge region

antigen

binding

sites

exposed surface

hidden surface
hidden surface
hinge region heavy chain carbohydrate light chain
hinge region
heavy chain
carbohydrate
light chain
sites exposed surface hidden surface hinge region heavy chain carbohydrate light chain Kappa target Lambda
sites exposed surface hidden surface hinge region heavy chain carbohydrate light chain Kappa target Lambda

Kappa

target
target
target
target

target

target
target
sites exposed surface hidden surface hinge region heavy chain carbohydrate light chain Kappa target Lambda

Lambda

Previously

hidden surface

and antibody

The Binding Site

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Free Light Chain Quantitation: Reference Intervals

 

Reference Range (95% interval)

Diagnostic Range (100% interval)

Kappa FLC

0.33-1.94 mg/dL

 

Lambda FLC

0.57-2.63 mg/dL

 

FLC k/l ratio

0.3-1.2

0.26-1.65

mg/dL   FLC k/l ratio 0.3-1.2 0.26-1.65 Katzmann, J. Clin Chem, 2002 Department of Laboratory
mg/dL   FLC k/l ratio 0.3-1.2 0.26-1.65 Katzmann, J. Clin Chem, 2002 Department of Laboratory

Katzmann, J. Clin Chem, 2002

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

100000 10000 Normals <70 yrs Normals >70 yrs 1000 Kappa LCMM Lambda 100 LCMM NSMM
100000
10000
Normals <70
yrs
Normals >70
yrs
1000
Kappa
LCMM
Lambda
100
LCMM
NSMM
10
Polyclonal
1
1
10
100
1000
10000
100000
Free light chain serum Lambda (mg/L)

Free light chain serum Kappa (mg/L)

serum Lambda (mg/L) Free light chain serum Kappa (mg/L) The Binding Site Department of Laboratory Medicine

The Binding Site

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Abnormal rFLC in Plasma Cell Disorders

Diagnosis

n

Abnormal rFLC, %

Multiple myeloma (MM)

Intact Ig MM “Non secretory” MM Light chain MM

MGUS

AL amyloidosis

LCDD

Smoldering MM

Plasmacytoma

1706

96

33

73

252

100

1262

33

467

95

28

93

345

90

116

47

Katzmann, J. Clin Chem, 2009 Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation
Katzmann, J. Clin Chem, 2009 Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation
Katzmann, J. Clin Chem, 2009 Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation
Katzmann, J. Clin Chem, 2009 Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation

Katzmann, J. Clin Chem, 2009

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Diagnostic Screening Panels: Sensitivity

Diagnosis (n)

Serum PEL/IFE/FLC 1 Urine PEL/IFE

Serum

Serum PEL/FLC

PEL/IFE/FLC

(all 5 assays)

(all 3 serum assays)

(2 serum assays)

MM 2 (467)

100%

100%

100%

WM (26)

100%

100%

100%

AL (581)

98.1%

97.1%

96.2%

SMM (191)

100%

100%

99.5%

MGUS (524)

100%

97.1%

88.7%

1 PEL=protein electrophoresis; IFE=immunofixation electrophoresis; FLC=quantitative free light chain

2 MM=multiple myeloma; WM=Waldenström's macroglobulinemia; AL=primary amyloidosis:

SMM=smoldering multiple myeloma; MGUS=monoclonal gammopathy of undetermined significance

MGUS=monoclonal gammopathy of undetermined significance Department of Laboratory Medicine and Pathology © 2010 Mayo
MGUS=monoclonal gammopathy of undetermined significance Department of Laboratory Medicine and Pathology © 2010 Mayo

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

IMWG Guidelines for Quantitative FLC: Impact on the Laboratory

Diagnosis:

 

Screening panel: serum PEL/IFE & FLC unless suspect AL.

 

Serum PEL & FLC is sufficient for initial screen

Prognosis:

& FLC is sufficient for initial screen Prognosis: Department of Laboratory Medicine and Pathology © 2010
& FLC is sufficient for initial screen Prognosis: Department of Laboratory Medicine and Pathology © 2010
& FLC is sufficient for initial screen Prognosis: Department of Laboratory Medicine and Pathology © 2010
& FLC is sufficient for initial screen Prognosis: Department of Laboratory Medicine and Pathology © 2010

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

A Long Term Study of Prognosis in MGUS

1384 patients in SE MN were identified with MGUS between 1960 and 1994. Patients were followed for an average of 8 years (11,009 total patient years).

115 progressed to MM, lymphoma, AL, Macroglobulinemia, CLL, or plasmacytoma

~1% of MGUS patients progress/year.

Initial size of M-spike and heavy chain isotype are prognostic for progression

and heavy chain isotype are prognostic for progression Kyle, et al, NEJM, 2002 Department of Laboratory
and heavy chain isotype are prognostic for progression Kyle, et al, NEJM, 2002 Department of Laboratory

Kyle, et al, NEJM, 2002

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

FLC and Progression in MGUS

Normal FLC ratio (K/L 0.26-1.65) Abnormal FLC ratio (K/L <0.26 or > 1.65) 0 5
Normal FLC ratio (K/L 0.26-1.65)
Abnormal FLC ratio (K/L <0.26 or > 1.65)
0
5
10
15
20
25
30
percent
500
10
20
30
40
60
0 5 10 15 20 25 30 percent 500 10 20 30 40 60 years Department

years

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Multivariate Analysis of Prognostic Factors for Progression of MGUS

Prognostic factor

Hazard ratio (95% C.I.)

p value

Abnormal FLC ratio

2.6 (1.7,4.2)

<0.001

Serum M protein size

2.4 (1.7,3.5)

<0.001

IgA, IgM, or biclonal IgA plus IgM

2.6 (1.7,4.0)

<0.001

or biclonal IgA plus IgM 2.6 (1.7,4.0) <0.001 Department of Laboratory Medicine and Pathology © 2010
or biclonal IgA plus IgM 2.6 (1.7,4.0) <0.001 Department of Laboratory Medicine and Pathology © 2010
or biclonal IgA plus IgM 2.6 (1.7,4.0) <0.001 Department of Laboratory Medicine and Pathology © 2010

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Prognosis of MGUS: Risk Stratification using M spike size, type and FLC ratio All 3
Prognosis of MGUS: Risk Stratification using M spike size, type and FLC ratio
All 3 factors abnormal
Any 2 factors abnormal
Any 1 factor abnormal
Serum M-spike <1.5 gm/dL, IgG Subtype and normal FLC ratio
0
5
10
15
20
25
30
Department of Laboratory Medicine and Pathology
Years
© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.
Percent
600
20
40

Risk Stratification Model Incorporating All 3 Predictive Factors

Risk Group

Number of

patients

Relative risk

Absolute

ROP at

20 years

20-year ROP after other causes of death

Low Risk

1. M protein <1.5 gm/dL,

2. IgG subtype,

3. normal FLC ratio

449

1

5%

2%

(0.1%/yr)

Low/Intermediate

Any 1 factor abnormal

420

5.4

21%

10%

High/Intermediate

Any 2 factors abnormal

226

10.1

37%

18%

High

All 3 factors abnormal

53

20.8

58%

27%

ROP: risk of progression

Rajkumar, SV, Blood,2005

58% 27% ROP: risk of progression Rajkumar, SV, Blood,2005 Department of Laboratory Medicine and Pathology ©

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Diagnostic Screening Panels: Sensitivity

Diagnosis (n)

Serum PEL/IFE/FLC 1 Urine PEL/IFE

Serum

Serum PEL/FLC

PEL/IFE/FLC

(all 5 assays)

(all 3 serum assays)

(2 serum assays)

MM 2 (467)

100%

100%

100%

WM (26)

100%

100%

100%

AL (581)

98.1%

97.1%

96.2%

SMM (191)

100%

100%

99.5%

MGUS (524)

100%

97.1%

88.7%

1 PEL=protein electrophoresis; IFE=immunofixation electrophoresis; FLC=quantitative free light chain

2 MM=multiple myeloma; WM=Waldenström's macroglobulinemia; AL=primary amyloidosis:

SMM=smoldering multiple myeloma; MGUS=monoclonal gammopathy of undetermined significance

MGUS=monoclonal gammopathy of undetermined significance Department of Laboratory Medicine and Pathology © 2010 Mayo
MGUS=monoclonal gammopathy of undetermined significance Department of Laboratory Medicine and Pathology © 2010 Mayo

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

IMWG Guidelines for quantitative FLC:

Impact on the Laboratory

Diagnosis:

Screening panel: serum PEL/IFE & FLC unless AL suspected.

Do we need IFE? Serum PEL & FLC are sufficient for MM screen.

Prognosis:

MGUS progression (rFLC, PEL, IFE)

Risk stratification should help define MGUS monitoring strategies

should help define MGUS monitoring strategies Department of Laboratory Medicine and Pathology © 2010 Mayo
should help define MGUS monitoring strategies Department of Laboratory Medicine and Pathology © 2010 Mayo
should help define MGUS monitoring strategies Department of Laboratory Medicine and Pathology © 2010 Mayo

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Prognosis Cut-points for FLC Ratio Depends on Plasma Cell Proliferative Disorder

Diagnosis

rFLC

MGUS

<0.26 or >1.65

Plasmacytoma

<0.26 or >4.0

Smoldering myeloma

<0.125 or >8

Symptomatic myeloma

<0.03 or >32

or >8 Symptomatic myeloma <0.03 or >32 Department of Laboratory Medicine and Pathology © 2010 Mayo
or >8 Symptomatic myeloma <0.03 or >32 Department of Laboratory Medicine and Pathology © 2010 Mayo
or >8 Symptomatic myeloma <0.03 or >32 Department of Laboratory Medicine and Pathology © 2010 Mayo

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

IMWG Guidelines for quantitative FLC:

Impact on the Laboratory

Diagnosis:

 

Screening panel: serum PEL/IFE & FLC unless AL suspected.

 

Do we need IFE? Serum PEL & FLC are sufficient for MM screen.

Prognosis:

 

FLC is prognostic in every PCD studied (MGUS, SMM, MM, AL, solitary plasmacytoma), and baseline values should be measured:

 

MGUS progression (rFLC, PEL, IFE)

 

Risk stratification should help define MGUS monitoring strategies

 

Smoldering myeloma progression (rFLC, PEL, %BMPC)

Symptomatic myeloma survival (rFLC, β2M, Alb)

Plasmacytoma survival (rFLC, IFE at 1 yr)

AL amyloidosis survival (dFLC)

Monitoring:

 
AL amyloidosis survival (dFLC) Monitoring:   Department of Laboratory Medicine and Pathology © 2010 Mayo
AL amyloidosis survival (dFLC) Monitoring:   Department of Laboratory Medicine and Pathology © 2010 Mayo

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Monoclonal IgG lambda protein

Monoclonal IgG lambda protein Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for Medical
Monoclonal IgG lambda protein Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for Medical
Monoclonal IgG lambda protein Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for Medical

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

CV of M-spike measurement vs. M-spike size

CV of M-spike measurement vs. M-spike size Department of Laboratory Medicine and Pathology © 2010 Mayo
CV of M-spike measurement vs. M-spike size Department of Laboratory Medicine and Pathology © 2010 Mayo
CV of M-spike measurement vs. M-spike size Department of Laboratory Medicine and Pathology © 2010 Mayo

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

CV of Monoclonal Protein Measurements in Stable Patients

 

Serum IgG

Measurable Serum M-spike

Measurable Urine M-spike

Measurable Serum iFLC

[IgG quant]

[M-spike>10g/L]

[M-spike>200

[iFLC>100

mg/24 hr]

mg/L]

CV

13.0%

8.1%

35.8%

28.4

(n=148)

(n=90)

(n=25)

(n=52)

35.8% 28.4 (n=148) (n=90) (n=25) (n=52) Katzmann, J. Clin Chem, 2011 Department of Laboratory
35.8% 28.4 (n=148) (n=90) (n=25) (n=52) Katzmann, J. Clin Chem, 2011 Department of Laboratory

Katzmann, J. Clin Chem, 2011

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Calculation of Biologic Variability

 

Serum M- spike (10 g/L)

Urine M- spike (200

Serum iFLC (100 mg/L)

mg/24hr)

Total CV

8.1%

35.8%

28.4%

Analytic CV

2.1%

4.5%

5.8%

Biologic CV

7.8%

35.5%

27.8%

4.5% 5.8% Biologic CV 7.8% 35.5% 27.8% Department of Laboratory Medicine and Pathology © 2010 Mayo
4.5% 5.8% Biologic CV 7.8% 35.5% 27.8% Department of Laboratory Medicine and Pathology © 2010 Mayo
4.5% 5.8% Biologic CV 7.8% 35.5% 27.8% Department of Laboratory Medicine and Pathology © 2010 Mayo

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Decrease needed in each assay for significance at various probability thresholds

Test

n

Total CV

P=0.05

P=0.90

P=0.95

Serum M- spike >10g/L

90

8.1%

7.4%

17.2%

20.1%

IgG

148

13.0%

11.7%

26.1%

30.3%

Urine M-spike >200 mg/24 hr.

25

35.8%

28.8%

56.5%

62.9%

iFLC

52

28.4%

23.7%

48.3%

54.5%

>100mg/dL

52 28.4% 23.7% 48.3% 54.5% >100mg/dL Department of Laboratory Medicine and Pathology © 2010 Mayo

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

IMWG Guidelines for quantitative FLC:

Impact on the Laboratory

Diagnosis:

 

Screening panel: serum PEL/IFE & FLC unless AL suspected.

 

Serum PEL & FLC are sufficient for MM screen.

Prognosis:

FLC is prognostic in every PCD studied (MGUS, SMM, MM, AL, solitary plasmacytoma), and baseline values should be measured.

 

Risk stratification should help define MGUS monitoring strategies

Monitoring:

Oligosecretory PCD (AL, NSMM, LCDD…) 50% change for partial response

Criteria may change such that urine M-spike or serum FLC can be used to monitor

90% reduction = partial response

be used to monitor › 90% reduction = partial response Department of Laboratory Medicine and Pathology

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Problems for electrophoretic diagnosis and monitoring of plasma cell dyscrasias

Free light chains: identification & monitoring

FLC quantitation

Monitoring very large M-spikes on agarose

Ig and/or HLC quantitation

Monitoring small M-proteins in γ or β fractions

Monitoring small M-proteins in γ or β fractions • HLC quantitation Broadly migrating M-proteins :

HLC quantitation

Broadly migrating M-proteins : identification & monitoring

HLC quantitation

: identification & monitoring • HLC quantitation Department of Laboratory Medicine and Pathology © 2010 Mayo

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Comparison of Agarose M-spikes and Ig Nephelometry Quantitation

of Agarose M-spikes and Ig Nephelometry Quantitation IgA IgG IgM Murray et al, Clin Chem 2009
IgA IgG IgM
IgA
IgG
IgM

Murray et al, Clin Chem 2009

Quantitation IgA IgG IgM Murray et al, Clin Chem 2009 Department of Laboratory Medicine and Pathology

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Large IgG M-proteins are Artificially Low by SPEP Resulting in High Albumin Quantitation

Low by SPEP Resulting in High Albumin Quantitation Department of Laboratory Medicine and Pathology © 2010
Low by SPEP Resulting in High Albumin Quantitation Department of Laboratory Medicine and Pathology © 2010

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Overestimation of IgM by Nephelometry

Overestimation of IgM by Nephelometry Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for
Overestimation of IgM by Nephelometry Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for
Overestimation of IgM by Nephelometry Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for
Overestimation of IgM by Nephelometry Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Quantitation of Large M-proteins

Biases exist between agarose electrophoresis

and nephelometry IgA has good agreement

IgG artificially lower results by SPEP

IgM artificially higher results by NEPH

Clinicians should be encouraged not to switch methods when monitoring M-protein.

couraged not to switch methods when monitoring M-protein. Department of Laboratory Medicine and Pathology © 2010
couraged not to switch methods when monitoring M-protein. Department of Laboratory Medicine and Pathology © 2010
couraged not to switch methods when monitoring M-protein. Department of Laboratory Medicine and Pathology © 2010

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

How to monitor Small M-proteins?

How to monitor Small M-proteins? Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for
How to monitor Small M-proteins? Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for
How to monitor Small M-proteins? Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Small monoclonal IgG kappa protein

Small monoclonal IgG kappa protein Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for
Small monoclonal IgG kappa protein Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for
Small monoclonal IgG kappa protein Department of Laboratory Medicine and Pathology © 2010 Mayo Foundation for

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Quantitation and monitoring of very small M-proteins: Fuzzy γ, elevated β or α

of very small M-proteins: Fuzzy γ , elevated β or α Department of Laboratory Medicine and
of very small M-proteins: Fuzzy γ , elevated β or α Department of Laboratory Medicine and
of very small M-proteins: Fuzzy γ , elevated β or α Department of Laboratory Medicine and

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Principle of Hevylite™ Assay

Principle of Hevylite™ Assay HLC junction Epitopes span the junction between immunoglobulin heavy chain and light

HLC junction

Epitopes span the junction between immunoglobulin heavy chain and light chain and therefore can quantitate each HL pair [GK, GL, AK, AL, MK, ML].

e can quantitate each HL pair [GK, GL, AK, AL, MK, ML]. The Binding Site Department
e can quantitate each HL pair [GK, GL, AK, AL, MK, ML]. The Binding Site Department

The Binding Site

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Potential of HL-pair Ratios for Monitoring Small M-proteins

MGUS

Number

Percent

Monoclonal

of cases

abnormal

protein

HLC-pair

isotype

ratios

IgG

726

45%

IgA

117

89%

IgM

156

90%

726 45% IgA 117 89% IgM 156 90% Department of Laboratory Medicine and Pathology © 2010
726 45% IgA 117 89% IgM 156 90% Department of Laboratory Medicine and Pathology © 2010
726 45% IgA 117 89% IgM 156 90% Department of Laboratory Medicine and Pathology © 2010

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

Identification of broadly migrating M-proteins:

Hevylite Reagents

Pretransplant August, 2006

M-proteins: Hevylite Reagents Pretransplant August, 2006 L K M A G PEL August 2010 IFE normal
M-proteins: Hevylite Reagents Pretransplant August, 2006 L K M A G PEL August 2010 IFE normal
M-proteins: Hevylite Reagents Pretransplant August, 2006 L K M A G PEL August 2010 IFE normal

L

K

M

A

G

PEL

August 2010

Pretransplant August, 2006 L K M A G PEL August 2010 IFE normal IgA 757 mg/dL

IFE normal

IgA 757 mg/dL

κ/λ 1.08

IgA κ = 638 mg/dL [55-292] IgA λ = 115 mg/dL [39-249]

AK/AL = 5.6 [0.7-2.2]

GK/GL = 1.8 [1.2-3.6]

Increase in IgA is not polyclonal - preferential expression of IgA κ

Donato et al, Clin Chem, 2011

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.

MADDOG:MADDOG: Myeloma,Myeloma, Amyloid,Amyloid, DysproteinemiaDysproteinemia DiseaseDisease OrientedOriented GroupGroup

Disease Disease Oriented Oriented Group Group Department of Laboratory Medicine and Pathology © 2010 Mayo
Disease Disease Oriented Oriented Group Group Department of Laboratory Medicine and Pathology © 2010 Mayo

Department of Laboratory Medicine and Pathology

© 2010 Mayo Foundation for Medical Education and Research. All Rights Reserved.