March 15 (LAB) Flashcards by John Lesther Alonsagay

Condensation method

Ortho-toluidine (dubowski method)

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Condesation of glucose with witha primary aromatic amine in glacial acetic acid, forming an equilipibrium mixture of a glycosylamine and the corresponding schiff base

Ortho-toluidine

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glycosylamine + schiff’s base

Ortho-toluidine

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Colormetric glucose oxidase method (saifer gerstenfield method)

Glucose oxidase method

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Polarographic glucose oxidase

Glucose oxidase method

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Whole blood is __ lower than serum or plasma

11%

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leukocytosis (increased WBCs) is up to __ mins

90 mins

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is present in the gray tube
which is used for glucose determination

Sodium fluoride

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inhibits the glycolytic enzyme
enolase which acts as an anticoagulant

Sodium fluoride

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There is also __ which inhibits the
glycolytic enzyme, glyceraldehyde-3-phosphate
dehydrogenase

acetate

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Glycolysis decreases serum glucose by approximately
__ (5 to 10 mg/L) in normal
uncentrifuged coagulated blood at room temperature

5% to 7% in 1 hour

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Refrigerated (___C)
○ Serum or Plasma: stable up to ___
○ Whole blood: 2mg Na fluoride per mL of whole
blood (48 hrs)

2-8*C, 48 hrs

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Refrigerated (___C)
○ Serum or Plasma: stable up to ___
○ Whole blood: 2mg Na fluoride per mL of whole
blood (48 hrs)

2-8*C, 48 hrs

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Different lab methods to determine glucose levels in the
sample

○ Chemical method
○ Enzymatic method

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A. Oxidation-Reduction Method
1. Alkaline Copper Reduction Method:

a. Folin Wu Method
b. Nelson Somogyi Method
c. Neocuproine Method
d. Benedict’s Method

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Alkaline Ferric Reduction Method

a. Hagedorn Jensen

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B. Condensation Method

a. Dubowski Method

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Principle: reduction of cupric ions to cuprous
ions forming cuprous oxide in hot alkaline
solution by glucose

Alkaline Copper Reduction Method

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The glucose reduces cupric ions present in the
alkaline copper reagent to cuprous ions or the
cupric sulfate is converted to cuprous oxide
which reduces the phosphomolybdic acid to
phosphomolybdous acid

Folin Wu Method

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Sugars with reducing properties are rising out
of the presence of the potential aldehyde or
keto group called the reducing sugars
● Ex of reducing sugars

Nelson Somogyi Method

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The reducing sugars when heated with the
alkaline copper tartrate reduce copper from the
cupric state and cuprous oxide is formed

Nelson Somogyi Method

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When cuprous oxide is heated with
arsenomolybdate, there will be reduction of ___ to ___
which has the color ___

molybdic acid to molybdenum, blue

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420 nm

Folin Wu method

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The copper in oxidation state reacts with
neocuproine forming a complex, this complex is
extracted into a chloroform methanol mixture
giving a yellow or yellow-orange solution

Neocuproine method

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Used for detection and quantitation of reducing substances in body fluids like blood and urine

Benedict's method

Can be used for identification of glucose in the urine

Benedict's method

It has a principle that when the reducing sugars are mixed with the Benedict’s reagent and heated, a reduction reaction causes the benedict’s reagent to change color

Benedic'ts method

Color varies from green to brick red, rusty brown = depending on the amount of sugar

Benedic'ts test

Aldolases are easily oxidized to yield carboxylic acid

Fehling's test

■ Benedict’s solution contains the Cu 2+ ___ ■ Fehling’s solution contains the Cu 2+ ___

citrate, tartrate

It involves reduction of yellow ferricyanide to a colorless ferrocyanide by glucose (inverse colorimetry)

Alkaline Ferric Reduction Method (Hagedorn Jensen)

Condensation of glucose with primary aromatic amine in glacial acetic acid, forming an equilibrium mixture of a glycosylamine and the corresponding Schiff base

Ortho-toluidine (Dubowski Method)

Acts on glucose but not on other sugars and reducing substances.

ENZYMATIC METHODS

is used to oxidize a dye compound to commonly chromogens are used in this type of method.

H2O2

is used to catalyze the second reaction.

Horseradish peroxidase

Increased levels of __,___,___ can cause falsely decreased values as a result of these substances being oxidized by peroxidase, which then prevents the oxidation and detection of the chromogen.

uric acid, bilirubin, and ascorbic acid

Measures rate of oxygen consumption which is proportional to glucose concentration.

Polarographic Glucose Oxidase

Glucose oxidase in the reagent catalyzes the oxidation of glucose by oxygen under first order conditions, forming ___.

hydrogen peroxide

Hydrogen peroxide is prevented from re-forming oxygen by adding ___,___,___, and ____.

molybdate, iodide, catalase and ethanol

___ can be used to perform the direct measurement of oxygen by polarographic technique.

Oxygen consumption

More accurate than glucose oxidase method

Hexokinase Method

Hexokinase in the presence of ATP converts glucose to ___

glucose 6 phosphate

___ is added to shorten the time

Mutarotose

___ is touched to a drop of blood and inserted into the meter which gives a digital reading of the blood sugar; gives the unit of mmol/L

Dextrostics

An enzyme-impregnated strip used with a small portable electronic colour-measuring device for convenient estimation of the blood sugar levels by diabetics.

Dextrostics

used for continuous monitoring of glucose levels in people with diabetes

Interstitial glucose monitoring device

Laboratory Testing for GLUCOSE

1. Random plasma glucose (random blood sugar) 2. Fasting plasma glucose (FBS) 3. Tolerance test 4. HbA1c (glycosylated hemoglobin test) 5. Fructosamine 6. Urine Microalbumin 7. Ketone testing

Random Plasma Glucose

● Formerly random blood sugar (RBS) ● Specimens collected anytime of the day ● Usually done in a glucometer ● NO NORMAL VALUES ○ Bcs the glucose is measured randomly

First step to measure the glucose in the blood is to convert glucose concentration into ___ or amount or the current signal

voltage

●___ To determine how well your body metabolizes glucose over a required period of time ● ___ samples in total

Oral glucose tolerance test, 3

Hemoglobin A is irreversibly glycosylated at one or both N-terminal valines of the B-chains of the tetrameric hemoglobin molecule (International Federation of Chemistry Working Group on HBA1c)

HbA1c

Is the largest subfraction of normal HBA involved in diabetic and nondiabetic individuals

HbA1c

It will cause the decreased value of your HBA1c

Hemoglibonpathy

A genetic defect that results in abnormal structure of one of the globin chains of the hemoglobin molecule

Hemoglibonpathy

Also known as the glycosylated albumin

Fructosamine

Most useful if the HBA1c is unreliable due to hemoglobinopathy or hemolysis

Fructosamine

Most widely used to assess short-term (__to__ week) glycemic control

Fructosamine, 3-6 weeks

Test to detect very small levels of protein (albumin) in urine

Urine Microalbumin

beta-HBA, acetoacetic acid, and acetone

Ketone

greatly increased in the Keto acidosis due to the altered rate and elevated levels of ___ in hepatic mitochondria

NADH

In serum acetone, it is indicative of defect in carbohydrate metabolism when the same acetone is increased

beta-HBA, acetoacetic acid, and acetone

Used of ferric chloride reacted with acetoacetic acid to produce a ___ color

Gerhatt's, Red color

■ Reacts with acetoacetic acid in an alkaline pH to form a ___ color ■ Urine reagent strip test and Acetest tablets

Sodium Nitroprusside, Purple Color

To detect either 3-b-hydroxybutyric acid or acetoacetic acid

3-hydroxybutyrate dehydrogenase

March 15 (LAB) Flashcards

(64 cards)