This document discusses monoclonal protein interferences in clinical chemistry assays. It presents a case study of a 58-year-old male with fatigue, weakness, and shortness of breath who was found to have extreme hyperphosphatemia. Further testing revealed an IgG-kappa monoclonal protein that was interfering with the phosphate assay, causing a pseudohyperphosphatemia. The document outlines various ways that monoclonal proteins can interfere with clinical chemistry assays, producing spurious results for analytes like bilirubin, HDL cholesterol, CRP, sodium, and total protein. It emphasizes the importance of laboratories identifying and investigating such interferences to avoid unnecessary testing or treatment of patients.

1. Monoclonal Protein Interferences
in Clinical Chemistry
Dr Sutirtha Chakraborty,
MD (Biochemistry), FACB (USA)
Chief Consultant , Dept. of Biochemistry
Peerless Hospital, Kolkata 1

2. CASE STUDY
• 58 year old male , known hypertensive on
treatment.
• Presents to hospital with progressive fatigue &
weakness for the past 2 months.
• Patient also complained of shortness of breath
for the past 2 weeks.
• Hb 9.5 gm/dL( RI: 12 – 15)
• Normocytic, Normochromic type
• WBC count(total & differential) –Normal
2

3. • Creatinine 2.05 mg/dl (RI 0.6 – 1.2 mg/dl)
• Urine Albumin(dipstick): Trace
• Serum Sodium 137 mmol/L (RI: 135-145)
• Serum Potassium 3.9 mmol/L (3.5 – 5.3)
Nephrologist Consult:
Serum Calcium, Phosphate and Uric Acid
Case Study 1…….Contd.
3

4. • Serum Calcium : 10 mg/dL (RI: 8.5 – 10.5)
• Serum Uric Acid: 9.0 mg/dL (RI: 3.5 – 7.0)
• Serum Phosphate: 28.4 mg/dl (RI: 2.5 – 4.5)
• Plasma PTH: 22 pg/mL (RI: 15 – 65)
•What could be the cause of extreme
hyperphosphatemia in this patient?
4

5. Spurious Lab Results
• Significant number of laboratory results and
reports are factitious or misinterpreted.
• Such laboratory results often lead to
unnecessary testing or treatment.
• An observant Laboratory Physician should
identify, intervene & investigate the error.
• Spurious/ Factitious results most commonly
occour due to assay Interference in the
“Analytical Phase”
• Major cause of reputational risk for labs.
5

6. Monoclonal Gammopathy
• Expansion of a single Ig-secreting plasma cell
population.
• Most cases involve IgG or IgA monoclonal cell
populations. About 15-20% are composed of IgM
monoclonal cells.
• Disorders associated with Monoclonal Protein:
MGUS, Multiple Myeloma (MM), Waldenström
macroglobulinemia (WM), amyloidosis (AL) or other
lymphoproliferative disorders.
6

7. Ig G-Kappa mediated Spurious
Hyperphosphatemia
• Loss of linearity
• ? Monoclonal Protein
• Serum Total Protein: 8.1 g/dL, Albumin 3.5 g/dL
• Serum Protein Electrophoresis : Sharp Band of
restricted mobility in Gamma region
Immunofixation:
Positive for Ig G - Kappa
7

8. 8

9. Interference as a result of Ig G – Kappa
monoclonal protein
1 2 3
Image: from Dr Graham Jones, Australia9

10. Pseudohyperphosphatemia
• The frequency of immunoglobulin-induced
laboratory errors is variable and probably
underreported.
• up to 1 in 4 patients with monoclonal
gammopathy show interference in PO4 assay.
• Pseudohypophosphatemia can also be seen
as a result of such interference.
• Various “wet chemistry” phosphomolybdate
assay is vulnerable to this interference.
10

11. Spurious Hyperbilirubinemia
• Monoclonal protein interferences occur with
total bilirubin measurements leading to false
high results.
• Results typically show T Bil > 15 mg/dL with
corresponding increased Indirect Bilirubin as
result of normal direct Bil.
• Apparently results mimic Hemolytic anemia.
• Common Culprit: Ig G-Lambda , Ig M (in WM)
11

12. Pantanwitz et al. Arch of Pathology 200412

13. Ig M interference with HDL Cholesterol Assay
Female, 58 years
Diagnosed with WM
Total C : 202 mg/dL
HDL C: 11 mg/dL
LDL C : 139 mg/dL
TG: 165 mg/dL
VLDL : 52 mg/dL
13

14. Spuriously Elevated CRP
• CRP > 300 mg/L
• Procalcitonin : 0.07
ng/mL
• Full Blood Counts:
Normal
• Blood Cultures: - Ve
• Final Diagnosis:
Light chain myeloma
14

15. Hypercalcemia & Myeloma
• Ig A Myeloma
• Extensive bone lytic lesions.
• BM Plasma Cells 30 %
• SPE : M band ( 4.8 g/dL)
• Serum Calcium 19.3 mg/dL
(Arsenazo Method)
To treat or not to treat this
hypercalcemia ?????????
15

16. Monoclonal Protein Interferences
Dalal et al. AJCP 2009
16

17. Strategies: What to do?
• Check the serum colour to see if it is truly icteric.
• Spectrophotometric measurement using an I
index, if available on the autoanalyzer. (LIH
index)
• Rerun the assay to demonstrate any imprecision
beyond what is typically observed.
• Check for loss of linearity (Serial dilutions)
• Measure the analyte using a different method.
• Always Correlate result with the clinical
information.
17

18. Multi-layer Thin Film Dry Slide Technology
18

19. Simple techniques for removing
Monoclonal Protein Interferences
1) Salting Out Methods
2) Precipitation with TCA
3) Ultrafiltration
4) Dialysis
5) Polyethylene Glycol (PEG)
Caution: Make sure that the analyte of interest is
insensitive to this procedure.
(Works well with Phosphate, Uric Acid, Bilirubin but not with
CRP, HDL-C)
19

20. 20

21. Pseudohyponatremia
• Caused by displacement
of serum water by
elevated concentrations
of serum lipids or
proteins.
• Indirect ISE involves
sample dilution and will
produce spuriously low
sodium.
• Occurs when TP > 12g/dL
Fortgens P, Archives of Path & Lab Med ,2011
21

22. Monoclonal protein interference in
the Preanalytical Phase
Chakraborty et al. CCLM 2014 22

23. 23

24. Which reagent do you choose ?
• Total Protein Regent 1
• Method: Biuret
• CV% - 1.8 %
• EQAS Outliers - None
• Pack Size – Same
• CPT: Rs 5/test
• Total Protein Regent 2
• Method: Biuret
• CV% - 1.9%
• EQAS Outliers - None
• Pack Size – Same
• CPT: Rs 1.25/test
24

25. Case Study
• A 50-year-old woman was being treated for
sudden-onset sensorineural deafness.
• LFT showed Total protein 10.1 g/dL and
albumin was 3.5 g/dL.
• Results Rechecked and released with a
comment “Marked A:G ratio reversal noted
advised Serum Protein Electrophoresis”
• Subsequent tests requested : SPE, IF, Sr. Calcium
25Chakraborty , Clin Chem 2015

26. • Normal Electrophoretic
pattern with IF negative.
• Dextran is used in the
management of sudden
hearing loss.
• Dextran Interference on
Biuret Assay?????
• So which reagent do we
choose?
26

27. JCLA, DECEMBER 2014
Conclusion:
“The results show that it is possible to use analytical interference
for diagnostic purposes, and most importantly, almost all cases
were identified at an early stage of the disease, when associated
clinical manifestations were not yet observed”.
27

28. Key Learning Points
• Since systemic deproteinisation of serum is no longer applied
in modern assays, interferences caused by monoclonal
proteins will continue to occur.
• All chemistry assays in patients with known monoclonal
gammopathies should be reviewed.
• Always check Manufacturer Kit Insert for possible
interferences.
Awareness is the key!
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29. Thank You
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