The document discusses Hemoglobin A1C (HbA1c) testing, which measures average blood sugar levels over the past 3 months. It summarizes that HbA1c is used to diagnose prediabetes and diabetes, and help manage diabetes treatment. The accuracy of HbA1c tests can be affected by hemoglobin variants or fetal hemoglobin levels. There are three main methods to measure HbA1c: chromatography based HPLC assay, antibody based immunoassay, and enzyme based enzymatic assay. HbA1c is not affected by short-term blood sugar fluctuations and does not require fasting for the test.

1. COLLEGE OF HEALTH SCIENCES AND MEDICINE
SCHOOL OF MEDICAL LABORATORY SCIENCE
DEPARTMENT OF MEDICAL LABORATORY SCIENCE
ASSESSMENT TYPES: SEMINAR PRESENTATION FOR 4TH YEAR MLS
STUDENTS
SEMINAR PRESENTATION TITLE: Hemoglobin A1C(HbA1c)
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2. GROUP MEMBERS
• Name ID Number
1.Melkamu Abebe...........................RU1971/12
2. Kuba Girma ……………………RU1862/12
3. Almaz Desta.............................….RU1191/12
4. Misgana Abay …………………..RU2024/12
5.Arersa Belema……………….........RU 1227/12
6. Gedefew betseha………………….RU1645/12
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3. Introduction
• Glucose is a type of sugar in the blood that comes from the foods we eat. Our cells
use glucose for energy. A hormone called insulin helps glucose get into cells.
• In diabetes the body doesn't make enough insulin, or the cells don't use it well. As
a result, glucose can't get into the cells, so the blood sugar levels increase.
• Hemoglobin A1C or HbA1c test is a simple blood test that measures average
blood sugar levels over the past 3 months.
• It's one of the commonly used tests to diagnose prediabetes and diabetes, and is
also the main test to help health care team to manage the diabetes.
• The A1C test is also called the glycated hemoglobin, glycosylated hemoglobin,
hemoglobin A1C or HbA1c test.
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4. Hemoglobin A1C or HbA1c test
• The sugar in the blood is called glucose. When glucose builds up in the blood, it binds to
the hemoglobin in the red blood cells.
• The A1c test measures how much glucose is bound.
• If the glucose levels have been high over recent weeks, the hemoglobin A1c test will be
higher.
Normal range of Hemoglobin A1c Test
• The normal range for the hemoglobin A1c level is between 4% and 5.6%.
• Hemoglobin A1c levels between 5.7% and 6.4% mean prediabetes and a higher chance of
getting diabetes.
• Levels of 6.5% or higher mean diabetes.
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5. Significance of HbA1c Test
The results of an A1C test can help Physicians or other health care provider to:
• Diagnose prediabetes. If prediabetes, it has higher risk of developing diabetes and
cardiovascular disease.
• Diagnose type 1 and type 2 diabetes. To confirm a diabetes diagnosis,
• Monitor diabetes treatment plan. The result of an initial A1C test also helps establish
baseline A1C level. The test is then repeated regularly to monitor diabetes treatment
plan.
 How Often the Need for HbA1c Test
How often the need for the test depends on several things, like:
• The type of diabetes
• The blood sugar control
• The treatment plan
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6. Cont ….
• HbA1c test is performed once a year in the case of prediabetes, which means
chance of developing diabetes.
• If type 2 diabetes HbA1c test is performed twice each year.
• If type 1 diabetes three or four times HbA1c test is may performed each year.
It may also need the test more often if the diabetes plan changes or if start a new
medicine
• It’s not a fasting test. It can be tested any time of the day, before or after eating.
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7. Conversion factors
• Equations used when A1C(mmol/mol) is entered: A1C(%) = (A1C(mmol/mol) /
10.929) + 2.15.
• Estimated average glucose(mg/dL) = (28.7 * A1C(%)) - 46.7.
• Average Plasma Blood Glucose (mg/dl) = (HbA1c * 35.6) - 77.3.
• Average Plasma Blood Glucose (mmol/L) = (HbA1c * 1.98) - 4.29.
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8.  A1C test results generally correspond with the following results of blood sugar
levels:
HbA1C level
Estimated average blood
sugar (glucose) level
6% 126 mg/dL (7 mmol/L)
7% 154 mg/dL (8.6 mmol/L)
8% 183 mg/dL (10.2 mmol/L)
9% 212 mg/dL (11.8 mmol/L)
10% 240 mg/dL (13.4 mmol/L)
11% 269 mg/dL (14.9 mmol/L)
12% 298 mg/dL (16.5 mmol/L)
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9. Limitations of the A1C test
Some factors may interfere with the accuracy of A1C test results.
These include:
• Pregnancy
• Recent or heavy blood loss
• Recent blood transfusion
• Conditions that result in insufficient red blood cells (anemias)
• Hemoglobin variants
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10. Cont ….
• The accuracy of HbA1c methods can be affected adversely by the presence of
hemoglobin (Hb) variants or elevated levels of fetal hemoglobin (HbF).
• The effect of each variant or elevated HbF must be examined with each specific
method.
• The most common Hb variants worldwide are HbS, HbE, HbC, and HbD. All of
these Hb variants have single amino acid substitutions in the Hb β chain.
• HbF is the major hemoglobin during intrauterine life; by the end of the first year,
HbF falls to values close to adult levels of approximately 1%.
• However, elevated HbF levels can occur in certain pathologic conditions or with
hereditary persistence of fetal hemoglobin.
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11. Methods used to measure HbA1c
• HbA1c is not affected by short-term fluctuations in blood glucose
concentrations,
• Blood can be drawn for HbA1c testing without regard to when food was
eaten.
There are 3 major HbA1c testing methods currently available to clinical
laboratories.
• Chromatography based HPLC assay
• Antibody based immunoassay
• Enzyme based enzymatic assay
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12. 1.Chromatography based HPLC assay
Test Principles
• Assay uses an HPLC instrument and ion exchange or affinity column to separate HbA1c
molecules from another hemoglobin molecules.
• Based on the ratio of HbA1c peak area to the total hemoglobin peak areas.
HPLC
• A chemical (electrical) charge is present on the molecule of HbA1c, and the
amount of the charge differs from the charges on the different components of
hemoglobin.
• The molecule of HbA1c has difference in size from the other components.
• HbA1c can be separated by charge and size from the other hemoglobinA
components in blood by a procedure known as high pressure (or performance)
liquid chromatography (HPLC).
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13. Cont ….
Advantages
• HbA1c overestimation leads to aggressive glucose management, resulting in
more frequent hypoglycemic episodes .
Drawbacks
• Altering the normal process of glycation of HbA to A1C.
• Causing an abnormal peak on chromatography, making estimation of A1C
unreliable.
• Making the red blood cell more prone to hemolysis, thereby decreasing the time
for glycosylation to occur and producing a falsely low A1C result.
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14. 2.Antibody based immunoassay
Test Principles
• A typical method uses a specific antibody (usually monoclonal) to the glucose and
the first 5 to 10 amino acids of the β-chain. This antibody is latex coated .
• The agglutinator reacts with the antibody to give a scattering of light and an
increase in absorbance.
• From this the amount of HbA1c is calculated, and the total hemoglobin can be
determined by measuring at or near the Soret absorption band of hemoglobin (410
- 420nm) or by Drabkins method (oxidation and conversion to
cyanmethemoglobin) at about 540nm, or using the alkali hematin assay.
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15. Cont …
Advantages
• Reduces the scattering of light and the absorbance
Drawbacks
• Time required to complete the analysis.
• Technical skills required for handling.
• High price of reagents.
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16. Enzyme based enzymatic assay
Test Principles
• Lysed blood samples are subjected to proteolytic digestion.
• Glycated valines are released and serve as substrate for fructosyl
valine oxidase.
• The produced hydrogen peroxide is measured using a horseradish
peroxidase-catalyzed reaction with a chromogen.
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17. Cont ….
Advantages
• Enzymatic assay proved to be a robust and reliable method for HbA1c
measurement suitable for routine practice in clinical chemistry laboratories.
• The assay is designed to report %HbA1c values directly without need for a
separate measurement of total hemoglobin and is not adversely affected by
interferences from common hemoglobin variants in samples.
Drawbacks
• A disadvantage of the enzymatic method is its relatively high cost
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18. Summary
• Hemoglobin A1C or HbA1c test is a simple blood test that measures average
blood sugar levels over the past 3 months.
• It's one of the commonly used tests to diagnose prediabetes and diabetes, and is
also the main test to help health care team to manage the diabetes.
• The accuracy of HbA1c methods can be affected adversely by the presence of
hemoglobin (Hb) variants or elevated levels of fetal hemoglobin (HbF).
• The effect of each variant or elevated HbF must be examined with each specific
method.
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19. • A1C test results are reported as a percentage. A higher A1C percentage corresponds
to higher average blood sugar levels.
• Results for a diagnosis are interpreted as :Below 5.7% is normal,5.7% to 6.4% is
diagnosed as prediabetes,6.5% or higher on two separate tests indicates diabetes.
• There are 3 major HbA1c testing methods currently available to clinical laboratories:
Chromatography based HPLC assay, Antibody based immunoassay and Enzyme
based enzymatic assay.
• HbA1c is not affected by short-term fluctuations in blood glucose concentrations,
example due to meals, blood can be drawn for HbA1c testing without regard to when
food was eaten.
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20. References
1.American Diabetes Association: “A1C Test,” “A1C and eAG.”
2. National Institute of Diabetes and Digestive and Kidney Diseases: “The A1C Test
and Diabetes.”
3.Ask MayoExpert. Type 2 diabetes mellitus (adult). Mayo Clinic; 2018.
4.Diabetes overview. National Institute of Diabetes and Digestive and Kidney
Diseases. https://www.niddk.nih.gov/health-information/diabetes/overview/all-
content. Accessed Dec. 4, 2020.
5. American Diabetes Association. Standards of medical care in diabetes. Diabetes
Care. 2008;31(Suppl.1): S12–S33. [PubMed] [Google Scholar]
6.William JR, Jang HH, Taewoo K (2014) Systems and methods for determining the
percentage of glycated hemoglobin.
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