3-Amylase
1. pancreatic amylase-pancreas 2.salivary amylase- salivary glands
a-amylase originates from what organ?
a-amylase
are hydrolytic enzymes which break down starch to maltose
The decrease in intensity of dark blue color at 660 nm. Rate of increasing absorbance at 405 nm: resulting to p-nitrophenol from hydrolysis
mode of measurement in amyloclastic & enzymatic/colorimetric
1cm
optical path
pancreas intestinal tract
pancreatic amylase is produced by __? and released into the ___
kidney urine urinary amylase activity
the amylase present in the body is eliminated through the ____ and excreted in ___ therefore elevation of serum activity is reflected in a rise of ??
U/L or units/100 ml sample U/L or µkat/L
unit of measurement in amyloclastic & enzymatic/colorimetric
200U/L max of 0.35 diluted 1+9 with NaCl multiply x 10
upper limit on automated systems of a-AMY upper limit on manual procedure? -if values exceed, what to do?
660nm Hg 405nm
wavelength used in amyloclastic & enzymatic/colorimetric
against reagent blank
measurement of a-AMY
Anhydrous disodium phosphate (pH 7.0 ± 0.1; Na2HPO4) Good's Buffer (pH - 7.15)
Buffer used & pH in amyloclastic & enzymatic/colorimetric
3U/L
lower limit of a-AMY
Endpoint Spectrophotometry UV, 3 point Kinetic, and Spectrophotometry
category of estimation in amyloclastic & enzymatic/colorimetric
Good's buffer (pH7.15; 0.1mol/L) NaCl (62.5mmol/L) Mg2Cl (12.5mmol/L) α-glucosidase (≥ 2kU/L) Good's buffer (pH7.15; 0.1mol/L) EPS-G7 (8.5mmol/L)
components of reagent 1 & 2 for a-amylase
Starch-iodine complex; Colorless (Initial color is Blue) P-nitrophenol (PNP; yellow if there is an increase in serum amylase activity; colorless if normal)
end product & color in amyloclastic & enzymatic/colorimetric
4554
factor used to calculate A/min of a-amylase
few hours after 12hrs after 5 days
in acute pancreatitis, blood amylase activity increases within how many hours from onset of abdominal pain, peaks and returns to normal
measurement of AMY in serum
it is used for the diagnosis of pancreatic disorders as well as detect devt of complications
-substrate 4,6-ethylidene-(G7)-p-nitrophenyl-(G1)-α-D-maltoheptaoside (EPS-G7) is cleaved by α-amylases into various fragments. -further hydrolyzed in a second step by α-glucosidase producing glucose and p-nitrophenol. -increase in Abs represents = the total (pancreatic and salivary) amylase activity in the sample.
principle of a-amylase determination
7.5 minutes (5 minutes pre-warming - to equilibrate mixture, correct lag phase) 5 minutes (2 minutes initial)
reaction time time in amyloclastic & enzymatic/colorimetric
serum/plasma -male & female= <100 (<1.67) urine -female= <447 (<7.45) -male= <491 (<8.18)
reference range of amylase in both serum/plasma & urine
160 U/100 mL or 111-296 U/L
reference range of amylase in iodometric/amyloclastic method
salivary glands saliva
salivary amylase is synthesized in the_____ and secreted into the _____
Serum (0.1 ml) Serum (0.02 ml or 20 µL)
sample and sample volume used in amyloclastic & enzymatic/colorimetric
ascorbic acid- >30mg/dl bilirubin- >40mg/dl hgb- >550mg/dl lipemia- >1000mg/dl triglycerides
sources of interference
lipase
specificity of a-amylase for pancreatic disorders is not very high -elevated lvls are measured alo in non-pancreatic diseases (parotitis & renal insufficiency) -therefore for confirmation of acute pancreatitis, we need another measurement of ___?
7 days at 20-25°C; 7 days at 4-8°C; 1 year at -20°C 2 days at 20-25°C; 10 days at 4-8°C; 3 weeks at -20°C
stability of a-AMY in serum/plasma stability of a-AMY in urine
6 months at 2-8°C; 4 weeks at 15-25°C
stability of reagent
Lintner's Soluble Starch 4,6-ethylidene-(G7)-p-nitrophenyl-(G1)- α Lintner's Soluble Starch -D-maltoheptaoside (EPS-G7
substrate used in amyloclastic & enzymatic/colorimetric
37C
temperature
