Analytical Methods usually use in detecting Diabetic Patiet

1. Analytical Methods of
Determining Diabetes
Mellitus
HANISHA ERICA P. VILLAESTER

2. A. Blood Test
 Sample Collection
 Venipuncture
 Capillary Blood by Skin Puncture
 Done when venipuncture cannot be performed or when reducing frequency of needle sticks
is desired and less painful

3. A.1 FASTING BLOOD GLUCOSE TEST
 Person to be tested should be on normal diet for at least 3 days prior to testing
 Fasting for 8 – 12 hours
 Plasma or serum is used instead of whole blood
 More readily measured on automated equipment
 Glucose concentration is 10 – 15% higher than in whole blood

4. A.1 FASTING BLOOD GLUCOSE
FASTING BLOOD GLUCOSE
< 110 mg/dL Normal
110 – 126 mg/dL Pre-Diabetes
> 126 mg/dL Diabetes

5. A.2.a ORAL GLUCOSE TOLERANCE TEST
 Fasting for 8 – 12 hours
 Patients requested to drink anhydrous glucose dissolved in 250 – 300mL water
 Adults: 75g anhydrous glucose
 Children: 1.75g/kg of body weight anhydrous glucose
 Blood tested regularly in 30 minutes interval
 In conditions of insulin defiency, blood glucose levels got elevated due to impaired
utilization of glucose
 Required for confirmation
 Glucose Tolerance Curve is then plotted

6. NORMAL GLUCOSE TOLERANCE CURVE

7. DIABETIC GLUCOSE TOLERANCE CURVE

8. NORMAL VS DIABETIC

9. A.2.b Intravenous Glucose Tolerance Test
 Undertaken for patients with malabsorption
 Patient is given 25g of glucose dissolved in 100mL distilled water as intravenous
injection within 5 minutes
 Blood samples drawn at 10 minutes interval for the next 2 hours
 Interpretation:
 Normal individuals, blood glucose level returns to normal within 60 minutes
 Diabetes mellitus – decline is slow

10. A.2.b Intravenous Glucose Tolerance Test
 Factors Affecting Glucose Tolerance Test
 Acute Infection
 Liver Disease
 Hyperthyroidism
 Hypothyroidism
 Starvation

11. A.2.c Mini or Modern GTT
 Two samples are collected
 Fasting (zero hour)
 2 Hour Post Glucose Load
Zero Hour After 2 Hours
Normal Person < 110 mg/dL < 140 mg/dL
Diabetic Person > 126 mg/dL > 200 mg/dL
Increase Glucose Tolerance 110 – 126 mg/dL 140 – 199 mg/dL

12. A.3 Post Prandial Blood Sugar
 Patient advised to have normal meal
 Blood collected 2 hours after meal
Glucose Level
Normal < 140 mg/dL
Pre-diabetic 140 – 200 mg/dL
Diabetic > 200 mg/dL

13. A.4 Random Blood Glucose
 Only required during emergency
 Diabetis Mellitus: > 200 mg/dL

14. A.5 HbA1c
 Test that measures amount of glycated haemoglobin in your blood
 Glycated haemoglobin is a substance in red blood cell that is formed when blood
sugar attaches to haemoglobin
 Generally reflect the state of glycemia over the preceding 8 – 12 weeks thereby
providing an improved method of assessing diabetic control

16. Analytical Methods of
Determining Diabetes
Mellitus
KEZIA AIJELETH SHAHAR CABANGAL

17. Self monitoring blood glucose (SMBG)
People who take insulin should regularly self monitor
blood glucose.
For people with non-insulin treated type 2 diabetes
testing is most useful if patients use the results to learn
and alter behaviour, or medication.
“...SMBG is most useful if patients use
the results to learn, as part of an overall
diabetes education package….”

18. Laboratory tests to prevent and delay
complications of diabetes
People with diabetes usually die from
macrovascular complications of their
diabetes; namely cardiovascular disease.
This is influenced by all of the commonly
recognised risk factors for cardiovascular
disease as well as glycaemic control.
Fasting lipid levels are measured three
monthly until stable and then 6 - 12
monthly thereafter.
It is important that management should be
individualised
Parameter Optimal value
Total
cholesterol
< 4 mmol/L
LDL cholesterol < 2.5 mmol/L
HDL cholesterol > 1 mmol/L
TC:HDL ratio < 4.5
Triglycerides < 1.7 mmol/L
HbA1C < 7 mmol/L

19. Diabetic renal disease
The best way of testing for diabetic renal disease is by urinary albumin:creatinine ratio
(ACR) and serum creatinine with estimated glomerular filtration rate (eGFR). These tests
are performed on everyone with diabetes at diagnosis and repeated at least annually –
more frequently if there is proteinuria, microalbuminuria or reduced eGFR.
Albumin:creatinine ratio
 ACR provides an estimate of daily urinary albumin excretion.
 Microalbuminuria cannot be detected on a conventional urinary protein dip stick.
 Microalbuminuria is urinary albumin excretion between 30 and 300 mg/day; above
300mg/day represents proteinuria.
 ACR is best measured in the laboratory using a first morning urine sample where
possible when the patient is well.
 An abnormal initial test requires confirmation by testing on two further occasions. If
at least one of these tests is positive microalbuminuria has been confirmed.

20. Renal testing in diabetes
*Non-diabetic renal disease is suspected when there is absence of diabetic retinopathy in a person
with renal disease, there are urinary abnormalities such as haematuria or casts, or when there is
renal disease without microalbuminuria or proteinuria.
ACR
mg/mmol
(confirmed)
eGFR
mL/min/1.
732
Risk Management
men < 2.5
women < 3.5
and > 60
2 - 4% per year
progress to
microalbuminuria.
Annual ACR and eGFR. Good diabetes &
BP management.
men ≥ 2.5
women ≥ 3.5
or < 60
One third progress to
overt nephropathy.
CVD risk doubled.
Review ACR and eGFR at each visit.
Intensive management of glycaemia
and CVD risk factors.
Use ACE inhibitor and low-dose aspirin.
Avoid nephrotoxic drugs.
Investigate if suspicious of causes other
than diabetes*
> 30 or < 30
Almost all proceed to
end stage renal
disease or die
prematurely of CVD.
Overt nephropathy
Refer specialist

21. Other tests
Testing of LFTs is recommended for people with diabetes:
 at diagnosis,
 at the start of antidiabetic drug therapy, and
 at any other time indicated by clinical judgement
Other laboratory tests
In patients with type 1 diabetes, intermittent checks for
other autoimmune conditions may be useful. This could
include testing for thyroid dysfunction or coeliac disease.

22. 22Urinary Glucose
 -Glucose can be detected in urine using the specific test strips that contain
glucose
 oxidase, peroxidase, and a chromagen.
 -Other carbohydrates using Benedict's and Febling's reagents.

23. 23Urinary Ketones
 -Acetone and acetoacetic acid can be detected in urine using the AcetesTM or
 KetostixTM systems.
 -These tablets or strips use nitroprusside (sodium nitroferricyanide) to detect
ketones.

24. 24Urinary Ketones
 -Because beta-hydroxybutyric acid lacks a ketone group is not detected by
this assay.
 -Quantitative assays for acetoacetate and beta-hydroxybutyric acid are
available using beta-hydroxybutyrate dehydrogenase and either NADH or
NAD.

25. 25Urinary Ketones
 -If NAD is used as the cofactor and the reaction is buffered at around pH
9.0, beta-hydroxyburyric acid is measured.
 -On the other hand, a separate reaction using NADH and buffered around
pH 7.0 would measure acetoacetic acid.

26. 26Microalbuminuria
 -Diabetes mellitus causes progressive changes to the kidneys and ultimately
results in diabetic renal nephropathy.
 -This complication progresses over a period of years and may be delayed by
aggressive glycaemic control.
 -An early sign that nephropathy is occurring is an increase in urinary albumin.

27. 27Microalbuminuria
 -Microalbumin measurements are useful to assist in diagnosis at an early
stage and
 prior to the development of proteinuria.
 -Microalbumin concentrations are between 20 to 300 mg/d.
 -Proteinuria is typically greater than 0.5 g/d.

28. 28Proteinuria in Diabetes
 - Many people excrete small quantities of protein in urine, typically around 10
 mg/day of mainly low molecular weight proteins such as albumin.
 -Some diabetic patients develop albumin excretion rates 30 µg/min this range
 classed as microalbuminuria.

29. 29
METHODS FOR THE DETERMINATION OF GLUCOSE
 The most used
 methods of
glucose analysis
employ the enzymes
glucose oxidase or
hexokinas.
 A) Glucose
Oxidase
 B) Hexokinase

30. 30
SELECTED METHODS FOR THE MEASUREMENTS OF GLYCATED HAEMOGLOBINS
InterferenceMeasurementMethod
Carbamyl Hb
HbF
Temperature-
sensitive
HbA1cCation exchange
HbA1cMonoclonal antibody
Glycated HbAffinity chromatography
Phenyl boronate matrix
Latex agglutination
Fluorescence quenching

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