American Association for Clinical Chemistry
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NACB - Scientific Shorts
NACB - Scientific Shorts (formerly NACB Blog)
By William Winter, MD
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The patient was a teenage African American female with a 2-week history of polyuria and polydipsia. There was no weight loss or nausea. She also presented with bilateral pain in her lower extremities and chest pain. During transit by EMS, her capillary glucose measured with a point-of-care device was 368 mg/dL.

 

These were her labs on admission:

 

 

Patient

Result

Reference Interval

Venous pH

7.26

7.3-7.4

Venous pCO2

23 mm Hg

42-53

Venous pO2

73 mm Hg

35-45

Venous HCO3-

10 meq/L

21-27

Venous base deficit

16 meq/L

0-3

Sodium

134 mmol/L

136-145

Potassium

4.1 mmol/L

3.3-5.1

Chloride

111 mmol/L

98-107

Carbon dioxide

8 mmol/L

22-30

Creatinine

0.62 mg/dL

0.40-1.10

BUN

6 mg/dL

6-20

Glucose

181 mg/dL

65-99

Urine glucose

>1000 mg/dL

Negative

Urine ketones

>150 mg/dL

Negative

 

More admission data:

 

 

Patient

Result

Reference Interval

WBC

20.50 K/uL

4.5-13.5K

Hemoglobin

9.9 g/dL

11.5-15.5

Hematocrit

28%

35-45

RBC count

3.16 million

4.0-5.2

MCV

87 fL

77-95

Red cell distribution width

16%

11-14

Corrected reticulocyte count

 

7.4%

0.5-1.8

Neutrophils

70%

40-80

Lymphocytes

18%

20-45

Monocytes

7%

2-10

Eosinophils

2%

0-8

Basophils

1%

0-2

Large unstained cells

3%

0-4

Platelet count

183 K/uL

150-450K

 

Hemoglobin A1c was requested and was measured at zero (0) percent. Below is the HPLC tracing. Why wasn’t any A1c measured?

 
Response:
This patient had been previously diagnosed with hemoglobin SC (i.e., heterozygosity for hemoglobin S and hemoglobin C). The clinical presentation of chest and leg pain is consistent with a sickling crisis including the acute chest syndrome probably precipitated by fluid loss and acidosis caused by new-onset type 1 diabetes with ketoacidosis.
 
The CBC displayed anemia and the peripheral smear reported moderate anisocytosis and sickle forms. In the absence of hemoglobin A0, hemoglobin A1 can not detected by HPLC. Hemoglobin A1c could be measured by immunoassay. However, with shortened red blood cell survival from hemolysis, the A1c measured by immunoassay could be falsely low. This would also be true if total glycohemoglobin were measured.
 
The HPLC tracing displayed 52% hemoglobin S and 39% hemoglobin C. Hemoglobin F was elevated at 5.4%. A hemoglobin electrophoresis from 2 years earlier revealed: 48% hemoglobin S, 7% hemoglobin F and 46% hemoglobin C.
 

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Posted by
On 1/30/2012

A patient with thse varients have no A0 peak so with ion exchange HPLC A1c can not be detected unless the blood is put on a HPLC with boronate affinity. Boronated affinity is interference free of variants to produce an accurate A1c.

Posted by
On 10/6/2011

why elevated Cl and decreased Na levels? This comment was approved by the NACBLOG editorial board. Please remember to add your name and affiliation!

Posted by
On 5/13/2011

Fructosomine: for individuals with hemoglobinopathies Hgb A1c is not an accurate method for monitoring diabetic control. For those patients with hemoglobinopathies fructosomine is an alternative , not for the last 12 week Hgb A1c level, but for atleast 2 -3 weeks. Better something than nothing at all. Fructosomine is not utilized by PCP's or endocrinologists because they are not aware of this test availablity. This comment was approved by the NACBLOG editorial board. Please remember to add your name and affiliation!

Posted by
On 5/11/2011

Something is causing the A1C to chelate out. Possible Sickle Cell patients. This comment was approved by the NACBLOG editorial board. Please remember to add your name and affiliation!

Posted by
On 5/2/2011

Several reasons may lead to false low or zero A1c values - especially on HPLC. From the chromatogram, it appears the analysis is done on Bio-Rad D10 HPLC in short mode. The method is based on principle of cation exchange chromatography and formal charge differences between hemoglobin variants. Although, it is highly rare that a whole blood sample would give a false result, but possibilities can not entirely ruled out that the alteration of the HPLC reagents (polarity or ionic nature of the mobile phase) or change in serum polarity cause false lowering of A1c. The inconsistency in polarity simply may have caused A1c to elute either early or late co-elution with other fractions. A clue can be found from the distribution of the overall peak area. Usually a zero A1c value is obtained when the actual A1c is already low in hypoglycemic condition. In this case, the reagents on HPLC should be changed, or calibration should be done. Also, I noticed the CO2 levels are low in the patient sample which may contribute to the blood polarity resulting in discrepancy in the normal chromatographic interaction leading to disappearance of A1c. In this case, immunoassay based analysis should provide proper values. Another possibility is also reported in literature is presence of other abnormal hemoglobin traits such as Hb-D, Hb-E, Hb-C or Hb-S. (Clin Chem, 2008, 130, 136 and Clin Chem, 2008, 54, 1277) Presence of these traits has different effects on the value of A1c (Hb-A1c). Usually modern HPLC based A1c analyzers have a longer mode which allows to detect this traits. If they are present, enzyme assay (not immunoassay) can be performed to obtain correct A1c (www.diazyme.com). Another possibility could be low lifespan of erythrocyte. Patient has slightly lower RBC and Hgb indices. This may contribute to lower A1c than normal. I hope this information will help to get some clue. This comment was approved by the NACBLOG editorial board. Please remember to add your name and affiliation!

About the Author
William Winter, MD
William Winter, MD 
 
Additional Resources
 
HbA1c: An Overview of Current Analytical Testing Issues. CLN February 2011

ADA Endorses HbA1c for Diabetes Diagnosis. CLN February 2010

Expert Committee Endorses HbA1c Test for Diagnosing Diabetes. CLN August 2009