This document discusses various laboratory tests used for the diagnosis and monitoring of diabetes, including the oral glucose tolerance test (OGTT), glycated hemoglobin (HbA1c), glycated albumin, fructosamine assay, and C-peptide assay. It provides details on each test, including what they measure, indications for use, limitations, and normal/abnormal ranges. It also outlines recommendations from diabetes organizations for screening and diagnosing both type 1 and type 2 diabetes using blood glucose criteria and HbA1c levels.

1. Diagnosis & Diagnostic
Tests of Diabetes
Dr Shahjada Selim
Associate Professor
Department of Endocrinology, BSMMU
Visiting Professor in Endocrinology, Texila American University, USA
Website: shahjadaselim.com

2. 1. Oral Glucose Tolerance Test (OGTT)
2. Glycated Hemoglobin (HbA1C)
3. Glycated Albumin
4. Fructosamine assay
5. Insulin Assay
6. Proinsulin Assay
7. C-Peptide Assay
Diagnostic Tests for Diabetes

3. Glucose Tolerance Test [GTT]
A glucose tolerance test is the
administration of glucose in a controlled
and defined environment to determine
how quickly it is cleared from the blood.
The test is usually used to test for
diabetes, insulin resistance, and
sometimes reactive hypoglycemia. The
glucose is most often given orally.

4. Types of GTT
 Standard Oral glucose tolerance test
 I/V Glucose tolerance test
 Mini Glucose tolerance test

5. Indication of OGTT
 In asymptomatic persons with sustained or
transient glycosuria.
 In persons with symptoms of diabetes but no
glycosuria or hyperglycemia.
 Persons with family history but no symptoms or
positive blood findings.
 In persons with or without symptoms of diabetes
mellitus showing one abnormal blood findings.
 In patients with neuropathies or retinopathies of
unknown origin.

6. Contraindications of glucose tolerance test
 There is no indication for doing GTT in
a person with confirmed diabetics
mellitus.
 GTT has no role in follow-up of
diabetics.
 The test should not be done in ill
patients.

7.  Patient should be on unrestricted carbohydrate
diet for 3 days.
 Patient should be ambulatory with normal
physical activity.
 Medications should be discontinued on the day of
testing.
 Exercise, smoking and tea or coffee are not
allowed during test period.
 OGTT carried out in the morning after patient has
fasted overnight for 8-12 hours.
Preparation of patient

8. • The OGTT is less accurate than the hyperinsulinemic-
euglycemic clamp technique (the "gold standard" for
measuring insulin resistance), or the insulin tolerance test,
but is technically less difficult. Neither of the two technically
demanding tests can be easily applied in a clinical setting or
used in epidemiological studies.
• HOMA-IR (homeostatic model assessment) is a convenient
way of measuring insulin resistance in normal subjects,
which can be used in epidemiological studies, but can give
Limitations of OGTT

9. Test
 A fasting venous blood
sample is collected in
the morning.
 Patients ingest 75 g of
anhydrous glucose in
250-300 ml of water
over 5 minutes. ( for
children, the dose is
1.75 g of glucose per
kg).

10. Test
In the classical procedures, the blood and
urine samples are collected at half hourly
interval of the next three hours.
A curve is plotted with the blood glucose
levels on the vertical axis against the time
of collection on the horizontal axis.
The curve so obtained is called glucose
tolerance curve.

11. Factors affecting GTT
a) Acute infections- Cortisol is secreted, the curve
is elevated and prolonged.
b) Hypothyroidism-A flat curve is obtained in
hypothyroidism. Thyroid hormone increases the
absorption of glucose from the gut.
c) Starvation- There is rise of counter regulatory
hormones, which show increased glucose
tolerance.

12. A1C ≥6.5% *
 Performed in a laboratory using a method that is
NGSP certified and standardized to the DCCT
assay - www.ngsp.org
 POC testing not recommended
 Greater convenience, preanalytical stability, and
less day-to-day perturbations than FPG and
OGTT
 Consider cost, age, race/ethnicity, anemia, etc.
American Diabetes Association Standards of Medical Care in Diabetes.
Classification and diagnosis of diabetes. Diabetes Care 2019; 39 (Suppl. 1): S13-S22

13. Glycated Hemoglobin
 Glycated hemoglobin
covers a number of
chemically different
modification resulting
from the non-enzymatic
and irreversibly binding of
different sugars to
different amino groups in
the hemoglobin molecule.
(Maillard Reaction )
Hemoglobin + glucose Aldimine Glycated hemoglobin

14. TYPE COMPONENTS
Glycated haemoglobin (Ghb) Haemoglobin in which glucose &/ any
other carbohydrates are bound to free
amino groups.
HbA1( fast haemoglobin) Carbohydrate bound to N- terminal of the
β- chain.
HbA₁а₁ Fructose-1, 6-biphosphate bound to the N-
terminal valine of the β-chain
HbA₁а₂ Glucose-6-phosphate bound to the N-
terminal valine of the β-chain
HbA₁ь Unknown carbohydrate residue bound to
the N-terminal valine of the β-chain
HbA₁с Glucose bound to the N-terminal valine
of the β-chain

15. Glycated hemoglobin
 HbA₁с gives information about the average
blood glucose concentration over a
retrospective period of time.
 Reflects the mean glucose concentration.
 Normally, less than 5% of hemoglobin is
glycated.

16. Glycated hemoglobin
 About 50% HbA₁с values results from the
blood glucose of the preceding 30 days , 40%
from the preceding 31- 90 days and only
10% from the period between the 91 – 120
days.
No effect of diet, exercise & insulin on test
results.
More informative.

17. Diagnostic Testing With 3 Different
Tests
Dealing with
Discordance
• Many people identified as
having diabetes using A1C
will not be identified as
having diabetes by traditional
glucose criteria, and vice
versa.
• When results of more than one test are available (amongst
FPG, A1C, 2hPG in a 75-g OGTT) and the results are
discordant, the test whose result is above diagnostic cut-point
should be repeated, and the diagnosis made on basis of the
repeat test.
FPG 2hPG
A1C

18. INDICATIONS
In all diabetes to monitor long term blood
glucose level control, index of diabetic
control:
7% HbA₁с – good
10% HbA₁с- Poorly ontrolled
> 13% HbA₁с- Very poor control.
To monitor patient compliance.

19. INDICATIONS
 To predict development & progression of
microvascular complication.
 For determining the therapeutic option
whether to use oral agents, insulin or β cell
transplantation.
 Also increasingly used for primary
diagnosis of DM.

20. At what interval should
HbA₁с be determined?
Treatment by time of
diabetes
Recommended
frequency
Type-1 DM (minimal
/conventional therapy)
4 times a year
Type – 1 DM (intensified
therapy)
Every (1) -2 months.
Type-2 DM Twice a year in stable
patients.

21. Glycated hemoglobin
High Values Low Values
Diabetes Mellitus Haemolysed specimen
PCOS Hereditary HbF
Hyperglycemia Neonate & Pregnancy
Glycosuria Fetal maternal
transfusion

22. Glycated hemoglobin
Falsely high values Falsely low values
Iron deficiency
anemia
Hemolytic anemia
Post spleenectomy Chronic blood loss
Alcohol poisoning Chronic Renal Failure
Lead toxicity Pregnancy

23. In conditions associated with an altered relationship
between A1C and glycemia, such as
sickle cell disease,
pregnancy (2nd and 3rd trimesters and the
postpartum period),
glucose-6-phosphate dehydrogenase deficiency,
HIV,
hemodialysis,
recent blood loss or transfusion, or erythropoietin
therapy,
only plasma blood glucose criteria should
be used to diagnose diabetes.
Do not Do HbA1c in…

24. HbA1C
 Blood sample can be drawn at any time of
day.
 HbA1c of 6 % corresponds to mean serum
glucose level of 7.5 mmol/L (135 mg/dl).
 With every rise of 1.0%, serum glucose
increases by 35 mg/dl.

25. Fructosamine assay
 Generic term for measurement of
all serum glycated protein though
the bulk being albumin.
 Does not appear to be influenced by
transient (stress) hyperglycemia.
 Unable to detect short term or
transient abnormalities in the blood
glucose concentration. Ex:
hypoglycemia.
 Reference range – in non diabetic-
2.4-3.4 mmol/l.
 Fructosamine / albumin ratio:- 54-
86 µmol/gm.

26. Fructosamine assay
Fructosamine test HbA1c
Measures average
blood glucose level
over the past two or
three weeks
Measures average
blood glucose level
over the past two
to three months.

27. Glycosylated Albumin
 Half -life of albumin is approximately 15
days.
 Glycated albumin level is believed to reflect
the glycemic change over a 2-week period.
 GA can be useful in evaluating the
therapeutic effect of recently substituted
hypoglycemic agents at an early stage.

28. Glycosylated Albumin
GA can also act as a valuable glycemic
control marker in diabetic patients with
various comorbidities since it is
unrelated to the metabolism of
hemoglobin.

29. Insulin assay
Measurement of
insulin level by
radioimmunoassay &
ELISA.
Crucial for type I DM.

30. Proinsulin Assay
 It is precursor molecule for insulin.
 Most proinsulin is converted to insulin and
C-Peptide, which are secreted in
equimolar amounts into the blood.
 The biological activity of proinsulin is only
about 10% of insulin, but the half life of
proinsulin is three times as long as insulin.

31. Proinsulin Assay
Elevated in:
 At onset of T1DM and in healthy sliblings of T1DM
patients.
 With established T2DM.
 Older patients.
 Pregnant .
 Obese pt with diabetes.
 Insulinomas.
 Functional hypoglycemia.
 Hyperinsulinemia.

32.  Released in circulation during
conversion of proinsulin to insulin
in equimolar quantities to insulin.
 Its level correlate with insulin level
in blood.
 Low C – peptide levels are
characteristic of type I DM.
 C-peptide levels are measured
instead of insulin levels because C-
peptide can assess a person’s own
insulin secretion even if they
receive insulin injections.

33.  The test may be used to help
determine the cause of hypoglycemia,
values will be low if a person has
taken an overdose of insulin but not
suppressed if hypoglycemia is due to
an insulinomas..
 Factitious hypoglycemia may occur
secondary to the surreptitious use of
insulin.
 Measuring C-peptide levels will help
differentiate a healthy patients from a
diabetes one.

34.  Blood glucose rather than A1C should be used to
diagnose T1DM in symptomatic individuals. E
 Screening for T1DM with an antibody panel is
recommended only in the setting of a clinical
research study or in a first-degree family
members of a proband with type 1 diabetes.
www.DiabetesTrialNet.org
Recommendations: T1DM
American Diabetes Association Standards of Medical Care in Diabetes.
Classification and diagnosis of diabetes. Diabetes Care 2020; 40 (Suppl. 1): S11-S24

35. Recommendations

36.  Screening for T2DM with an informal assessment of
risk factors or validated tools should be considered
in asymptomatic adults.
 Consider testing in asymptomatic adults of any age
with BMI ≥25 kg/m2 or ≥23 kg/m2 in Asian
Americans who have 1 or more additional DM risk
factors.
 For all patients, testing should begin at age 45 years.
 If tests are normal, repeat testing carried out at a
minimum of 3-year intervals is reasonable.
Recommendations: Testing for Type 2 Diabetes
American Diabetes Association Standards of Medical Care in Diabetes.
Classification and diagnosis of diabetes. Diabetes Care 2020; 40 (Suppl. 1): S11-S24

37. FPG, 2-h PG after 75-g OGTT, and the A1C
are equally appropriate.
In patients with diabetes, identify and, if
appropriate, treat other CVD risk factors.
Consider testing for T2DM in
overweight/obese children and adolescents
with 2 or more additional diabetes risk
factors.
Screening for Type 2 Diabetes (2)
American Diabetes Association Standards of Medical Care in Diabetes.
Classification and diagnosis of diabetes. Diabetes Care 2020; 40 (Suppl. 1): S11-S24

38. Fasting plasma glucose (FPG) ≥ 7.0 mmol/dL
(126 mg/dL)
OR
2-h plasma glucose ≥ 11.1 mmol/L (200 mg/dL)
during an OGTT
OR
A1C ≥6.5%
OR
Classic diabetes symptoms + random plasma
glucose ≥11.1 mmol/L (200 mg/dL)
Criteria for the Diagnosis of Diabetes
American Diabetes Association Standards of Medical Care in Diabetes.
Classification and diagnosis of diabetes. Diabetes Care 2020; 40 (Suppl. 1): S11-S24

39. Testing should begin at age 45 for all
patients, particularly those who are
overweight or obese.
Recommendations: Prediabetes
American Diabetes Association Standards of Medical Care in Diabetes.
Classification and diagnosis of diabetes. Diabetes Care 2019; 39 (Suppl. 1): S13-S22

40. FPG, 2-h PG after 75-g OGTT, and A1C, are
equally appropriate for prediabetes testing.
In patients with prediabetes, identify and,
if appropriate, treat other CVD risk factors.
Consider prediabetes testing in
overweight/obese children and adolescents
with 2 or more additional diabetes risk
factors.
Recommendations: Prediabetes (2)
American Diabetes Association Standards of Medical Care in Diabetes.
Classification and diagnosis of diabetes. Diabetes Care 2019; 39 (Suppl. 1): S13-S22

41. FPG 5.6–6.9 mmol/L
100–125 mg/dL
: IFG
OR
2-h plasma glucose 7.8–11.0 mmol/L
(140–199 mg/dL): IGT
OR
A1C 5.7–6.4%
Prediabetes*
* For all three tests, risk is continuous, extending below the
lower limit of a range and becoming disproportionately
greater at higher ends of the range.
American Diabetes Association Standards of Medical Care in Diabetes.
Classification and diagnosis of diabetes. Diabetes Care 2020; 39 (Suppl. 1): S13-S22

42. Blood glucose rather than A1C
should be used to dx type 1
diabetes in symptomatic
individuals.
Recommendation: Screening
for Type 1 Diabetes
American Diabetes Association Standards of Medical Care in Diabetes.
Classification and diagnosis of diabetes. Diabetes Care 2020; 39 (Suppl. 1): S13-S22

43. For all people, testing should be begin at
age 45 years. B
 If tests are normal, repeat testing carried
out at a minimum of 3-year intervals is
reasonable. C
Recommendations: Screening
for Type 2 Diabetes (2)
American Diabetes Association Standards of Medical Care in Diabetes.
Classification and diagnosis of diabetes. Diabetes Care 2019; 39 (Suppl. 1): S13-S22

44. Laboratory test for screening
Recommended screening test is fasting
plasma glucose.
American Diabetes Association recommends
screening for Type 2 DM in all asymptomatic
individuals ≥ 45 yrs of age using fasting
plasma glucose.

45. Laboratory test for screening
If fasting test is normal, screening test
should be repeated every three years.
If fasting blood glucose level is normal
but there is strong clinical suspicion then
OGTT/HbA1C.

46. Selective screening
 High risk individuals ---Obese
Family h/o DM
Hypertension
Dyslipidemia
Impaired glucose tolerance
Screening test is performed at earlier age ( 30 yrs ) and repeated
more frequently

47. Monitoring of Blood Glucose
SMBG

48. Self Monitoring of Blood Glucose- SMBG
Regular use of SMBG
devices by diabetic patients
has improved the
management of DM.
SMBG devices measure
capillary whole blood
glucose obtained by finger
prick and use test strips
that incorporate glucose
oxidase or hexokinase.

49. Self Monitoring of Blood Glucose- SMBG
SMBG devices yield
unreliable results at very
high and very low glucose
levels.
It is necessary to
periodically check the
performance of glucometer
by measuring parallel
venous plasma glucose in
the laboratory.

50. Battelino T, et al. Diabetes Care. 2019 Aug;42(8):1593-1603, Advani A. Diabetologia. 2020;63(2):242–252.
‘Time-In-Range’ & What’s Above/ Below From ‘In-Range’?
Target in glucose range
Below target glucose range
Above target glucose range
At least 17 hours
a day, more than
70% of their time
In a blood
glucose range of
70–180 mg/dL
People with diabetes should spent
Minimize
>17 hours
<1 hour
Level 2
hyperglycemia
Level 1
hyperglycemia
Level 1 hypoglycemia
Level 2 hypoglycemia
Time-In-Range
(>13.1 mmol/L)
(10-13.1 mmol/L)
(3.9-10 mmol/L)
(3.9-10 mmol/L)
(>13.1 mmol/L)
(10-13.1 mmol/L)
(<3.1 mmol/L)
(3.1-3.9 mmol/L)

51. HbA1c:Glycated hemoglobin.
Going Beyond A1c – One Outcome Can’t Do It All. Available at: https://diatribe.org/BeyondA1c. Accessed on: 31 January 2020.
Why Is Time-in–Range Important?
Patients experience different energy levels,
moods, QoL when they are ‘in-range’
vs. ‘out-of–range.’
Time-in–range can capture these
differences in a way A1C cannot
Patients at same A1C levels can
have different TIR profiles
Glycemic Variability?

53. SS