Amino Acida & Protein II Flashcards
is the reference method of determination
kjeldahl technique
not routinely used because it is TEDIOUS
kjedahl
in kjeldahl, the serum used should be pre treated with ___ to produce a____
tungstic acid, proteins free filtrate
colorimetric, is the most widely used, in fact it is also recommended by the IFCC panel
biuret method
principle of biuret method
formation of colored chelate between cupric ions and peptide bonds which forms a color violet
in biuret method there is a formation of colored chelate between ____ and ___ which forms a color violet
cupric ions and peptide bonds
the change of color in biuret method can be measured by the
spectrophotometer under 545 nm
also called lowry method, main reagent is a phenol reagent
folin ciocalteu method
principle of folin ciocalteu method
there is an oxidation of phenolic compound that will give a deep blue color
the absorbance of protein
UV absorption
UV absorption is the absorbance of protein at ____ due to the absorbance of the peptide bond’s _____
210-280 nm, specific wavelengths
there is a migration of charged particles in the electronic field
serum protein electrophoresis (SPE)
in SPE proteins will migrate from ____ to ____
negative to positive
in their migration ____ is the fastest
albumin
reference value for albumin
53-65%
second fastest to migrate
alpha 1 (2.5-5)
third fastest to migrate
alpha-2 (7-13%)
fourth fastest to migrate
beta (8-14% )
slowest to migrate
gamma (12-22%)
based on the refractive based on the proteins
refractometry
uses chemicals, SSA, determination of protein in the urine which uses acid which causes protein to coagulate, it also scatters light or blocks the light
turbidimetric and nephelometric methods
uses sodium sulfate salt
salt fractionation
salt fractionation will cause the ___ to be separated from the ___ (salting out)
globulin, albumin
most abundant protein in the blood
albumin
album is ____ to the severity of the liver disease
inversely proportional
albumin levels will decline if there are severe hepatocellular disease more than ____
3 weeks
functions of albumin
antioxidant, major buffer, transport
clinical adult range of albumin
3.5-5.0 g/dL
considered value for hypoalbuminemia
<3.5
value considered as hyperalbuminemia
> 5.0
- wherein nglobulins are precipitated in high salt concentration; albumin in supernatant is quantitated by biuret reaction
- labor intensive
salt precipitation
- nonspecific for albumin
- albumin binds to dye; caused shift in absorption maximum
methyl orange
- albumin binds to dye; causes shift in absorption
- many interferences
HABA (2,4- hydroxyazobenzene benzoic acid)
interferences of HABA
salicylates, bilirubin
- sensitive; overestimates low albumin levels;
- most commonly used dye
BCG (bromescol green)
- specific, sensitive , precise
- best
BCP (bromescol purple)
- accurate; gives overview of relative changes in different protein fractions
electrophoresis
proteins separated based on electric charge
electrophoresis
carrier of some hormones, lipids, metals, and antibodies
globulins
high globulin levels may be seen in
chronic infections, liver disease, rheumatoid arthritis, myelomas and lupus
lower globulin levels may be seen in
immunocompromised patients, poor dietary habits, malabsorption, liver and kidney disease
clinical adult range of globulins
2.0-4.0 g/dL
common causes of globulin increase
hypochlorydia (stomach doesnt produce acid anymore) liver disease (infection)
common causes of globulin decrease
anemia, hemorrhage
an important indicator of disease states
albumin/globulin ratio
inverted albumin/globulin ratio is seen in
cirrhosis, multiple myeloma, waldenstrom’s macroglobulinemia
albumin/globulin ratio clinical adult range
1.3 - 3.1
low albumin ratio suggests
ulcerative colitis, burns, and kidney disease
if ratio is ___ it could be a sign of disease to the liver, kidney, or the intestines, low thyroid activity, or leukemia
> 3
how to calculate for G (globulin)
TP-ALB
Ratio
ALB/G
