[LAB] LDH Flashcards
REFERENCE METHOD OF LDH
KINETIC METHOD BY BUHL AND JACKSON
MODIFIED BY WACKER
MAJOR ORGANS WHERE LDH IS DISTRIBUTED IN
HEART
LIVER
MUSCLE
KIDNEY
ELEVATED LEVELS OF SERUM LDH IS FOUND IN WHAT CLINICAL CONIDITONS
MI
LIVER DISEASE
RENAL DISEASE
ANEMIA
MALIGNANT DISEASES
PROGRESSIVE MUSCLE DYSTROPHY
IN WHAT DIRECTION CAN THE LD ENZYME BE MEASURED
FORWARD
REVERSE
[ADVANTAGES]
LACTATE TO PYRUVATE METHOD
BETTER REAGENT STABILITY
ELIMINATION OF PRE-INCUBATION
LINEAR RATE OF REACTION
LESS CONTAMINATION OF NAD
PRINCIPLE OF LDH
L-LACTATE + NAD
<—LDH (PH 8.3 TO 9.0) —>
PYRUVATE + NADH + H
COENZYME OF LDH
NAD
DOES LDH ACT ON D-LACTATE
NO
WHAT HAPPENS TO PYRUVATE AFTER FORMATION
PROCEED TO THE EMBDEN-MEYERHOFF PATHWAY
FOR THE GENERATION OF ENERGY
SOLUTIONS MIXED WITH NADH IN THE WACKER COLORIMETRIC METHOD
PHENAZINE METHOSULFATE
NITROBLUE TETRAZOLIUM
END PRODUCT OF THE WACKER COLORIMETRIC METHOD
BLUE COLORED COMPOUND
RELATIONSHIP OF THE ABS TO AMOUNT OF NADH AND LD ACTIVITY IN THE WACKER COLORIMETRIC METHOD
DIECTLY PROPORTIONAL
[DIRECTION OF REACTION]
COLORIMETRIC METHOD BASED ON WACKER
LACTATE TO PYRUVATE
[DIRECTION OF REACTION]
WROBLEWSKI AND LA DUE
PYRUVATE TO LACTATE
PH OF THE REACTION IN THE WROBLEWSKI AND LA DUE METHOD
PH 7.4 TO 7.8
RELATIONSHIP OF NADH BOLUME ANDNLD ACTIVITY IN THE WROBLEWSKI AND LA DUE METHOD
INDIRECTLY PROPORTIONAL
DIFFERENCE IN THE SPEED OF REACTION IN THE FORWARD AND REVERSE REACTION IN THE WROBLEWSKI AND LA DUE METHOD
REVERSE IS THREE TIMES FASTER
[ADVANTAGES]
WROBLEWSKI AND LA DUE METHOD
SMALLER SAMPLE VOLUMES
SHORTER REACTION TIME
[DISADVANTAGES]
WROBLEWSKI AND LA DUE METHOD
SUSCEPTIBLE TO SUBSTRATE EXHAUSTION
SUSCEPTIBLE TO LOSS OF LINEARITY
METHODS FOR SEPARATION AND QUANTITATION OF LDH ISOENZYMES
ELECTROPHORESIS
COLUMN CHROMATOGRAPHY
IMMUNO-INHIBITION
BABSON’S METHOD
KAPLAN AND BERER
USE OF ALPHA HYDROXYBUTYRATE AS SUBSTRATE
ELECTROPHORETIC MIGRATION OF THE LD ISOENZYMES
FASTEST — LDH1
SLOWEST — LDH6
BETWEEN LDH3/4 — LDH BOUND TO IMMUNOGLOBULIN (IgA/G)
LDH ISOENZYME ISOLATED IN THE COLUMN CHROMATOGRAPHY METHOD
LDH1
LDH2
METHOD USED FOR THE SCREENING OF AMI
COLUMN CHROMATOGRAPHY
LDH ISOENZYME ISOLATED IN THE IMMUNO-INHIBITION METHOD
LDH1
[PRINCIPLE]
IMMUNO-INHIBITION METHOD
AB TO THE M SUB-UNIT OF LDH IS ADDED TO THE SERUM SAMPLE
ANY ISOENZYME CONTAINING AN M-SUBUNIT WILL BE RENDERED INACTIVE
ONLY LDH1 DOES NOT HAVE THE M-SUBUNIT
LDH SUBUNIT THAT DOES NOT CONTAIN THE M-SUBUNIT
LDH1
LDH ISOENZYMES MEASURED IN BABSON’S METHOD
LDH1 TO LDH5 RATIO
SOLUTION USED TO INHIBIT LDH ISOENZYMES IN BABSON’S METHOD
M-LACTATE — LDH1
M-LACTATE-1M UREA — LDH5
LDH ISOENZYME MEASURED IN THE KAPLAN AND BERGER METHOD
LDH1
[PRINCIPLE]
KAPLAN AND BERGER
DETERMINATION OF LDH1 BY USING THE RATE OF HEAT PRODUCTION IN A BATCH TYPE OF COLORIMETER
LDH SUBUNIT WITH THE GREATER AFFINITY FOR ALPHA HYDROXYBUTYRATE
H SUBUNIT HAS A GREATER AFFINITY THAN THE M SUBUNIT
LD ISOENZYME MEASURED BY THE ALPHA HYDROXYBUTYRATE METHOD
LD1
[DISADVANTAGE]
APLHA HYDROXYBUTYRATE METHOD
NOT SPECIFIC TO LD1
LD2,3,4 ALSO HAVE VARYING AMOUNTS OF THE H SUBUNIT
EFFECT OF HEMOLYZED SPECIMENS ON LDH ACTIVITY
FALSE INCREASE
RBCs contain LDH activity 100-150x more than serum
DIFFERENCE OF SERUM AND RBC LDH ACTIVITY
RBCS HAVE LDH ACTIVITY 100-150X MORE THAN SERUM
EFFECT OF NOT ANALYZING THE SPECIMEN WITHIN 2 HOURS OF COLLECTION ON LDH ACTIVITY
FALSE DECREASE
LDH is stable for 2-3 days at RT
WHEN DOES THE LOSS OF LDH ACTIVITY INTENSIFY IN SAMPLES
4C > 25C
STABILITY OF LDH ACTIVITY IN SERUM
2-3 DAYS AT RT
TIME LIMIT FOR THE ANALYZING OF LDH SAMPLES UPON COLLECTION
WITHIN 2 HOURS OF COLLECTION
COLD LABILE LDH ISOENZYME
LIVER LDH
EFFECT OF FROZEN AND THAWED SPECIMENS ON LDH ACTIVITY
FALSE DECREASE
Liver LDH is cold labile and will be destroyed when frozen or thawed
EFFECT OF EDTA ON LDH ACTIVITY
FALSE DECREASE
EDTA chelates zinc (activator)
LDH ACTIVATOR ION
ZINC
EFFECT OF PLASMA SPECIMENS ON LDH ACTIVITY
FALSE INCREASE
Contamination with platelets which have LDH activity (LD3)
