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Lab ops > Electrophoresis > Flashcards

Electrophoresis Flashcards

1
Q

Electrophoresis definition

A

defined as the movement or
migration of charged particles in an electric
field

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2
Q

Electrophoresis is used to

A

Routinely used to separate serum proteins,
hemoglobin, enzymes, lipoproteins,
immunoglobulins, urine proteins, CSF
proteins, and other body fluid proteins

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3
Q

Electrophoresis is responsible in advances in what

A

Most of the remarkable advances in molecular
biology over the past few decades would have
been impossible without electrophoretic methods

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4
Q

Types of electrophoresis

A

Free electrophoresis and Zone electrophoresis

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5
Q

Free electrophoresis types

A

-Micro
Electrophoresis
-Moving
Boundary

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6
Q

Zone electrophoresis types

A

-Paper
Electrophoresis
Cellulose
-Acetate
Electrophoresis
-Gel
Electrophoresis

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7
Q

Iontophoresis is

A

he migration of small ions

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8
Q

Zone electrophoresis is

A

migration of charged
macromolecules in a porous support medium.

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9
Q

Electrophoresis system components

A

§ Charged particles
§ Support medium
§ Buffer
§ Electrical Power Source
§ Detecting system

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10
Q

Proteins are what and contain what

A

Proteins are amphoteric, containing both acidic and basic groups

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11
Q

Proteins charge is positive at

A

Low pH

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12
Q

Proteins zero (isoelectric)

A

at a particular higher pH

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13
Q

Proteins negative at

A

Alkaline pH

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14
Q

Electrophoretic mobility is proportional to

The effective mobility of a protein is

A

§ Electrophoretic mobility is directly proportional to the size of the
charge of the particle
§ The effective mobility of a protein is a function of pH

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15
Q

Support media types

A

§ AGAROSE GEL—most often used in clinical settings
§ POLYACRYLAMIDE GEL
§ STARCH GEL
§ CELLULOSE ACETATE
§ PAPER

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16
Q

Buffer functions

A

§ The buffer serves several functions:
§ Carries the applied current
§ Establishes and maintains a constant pH
§ Determines the electrical charge on the solute

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17
Q

Buffers

Protein separator buffers

A

§ Barbital buffers
§ Tris-boric acid-EDTA

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18
Q

Nucleic acid separation buffers

A

§ Buffers containing EDTA and Tris-acetate, Tris-borate, or Tris-
phosphate

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19
Q

Electrophoresis is conducted for a

A

§ Electrophoresis is conducted for a determined length of time
under conditions of either constant voltage or constant current.

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20
Q

Separated fractions are

A

§ Separated fractions are stained for visualization and densitometry

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21
Q

Numerous stains have been used and depend on

A

§ Numerous stains have been used, dependent upon the particular
analyte

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22
Q

Protein stains

A

§ Proteins:
§ Ponseau S Silver stain
§ Bromcresol blue Stains All
§ Light green SF Amido Black 10B
§ Coumassie blue R250 Colloidal Gold

23
Q

Lipoprotein stain

A

§ Sudan black B
§ Oil Red O
§ Coumassie brilliant blue R25

24
Q

Glycoprotein stains

A

§ PAS (Periodic-acid-Schiff)
§ Stains-All

25
Q

Nucleic acid stains

A

§ Stains All
§ Silver stains
§ Ethidium bromide
§ TOTO-1

26
Q

Enzyme stains

A

§ NADH (fluorescence)
§ Nitroblue tetrazolium chloride

27
Q

Esterase stains

A

§ Beta-naphthyl esters and tetrazotized o-dianisidine

28
Q

§ Cholinesterases & Phosphatase stains

A

§ 1-naphthyl phosphate and fast blue B

29
Q

Factors affecting rate of migration

A

§ Net charge of the molecule
§ Size and shape of the molecule
§ Strength of the electric field
§ Temperature
§ Ionic strength of buffer
§ Wick flow
§ Electroendosmosis/ (Endosmosis)

30
Q

Reference range of total protein

A

6.0-8.0g/dL

31
Q

Reference range of Albumin

A

3.5-5.0g/dL

32
Q

Reference range of a1-globulins

A

0.1-0.4g/dL

33
Q

Reference range of a2-globulins

A

.4-1.3g/dL

34
Q

B-globulin reference range

A

.6-1.3g/dL

35
Q

Reference range of y-globulin

A

.6-1.5g/dL

36
Q

The acute inflammatory response is

A
  • Immediate response occurs
    with stress or inflammation
    caused by infection, injury or
    surgical trauma
37
Q

Acute inflammatory response effects what values

A
  • normal or ↓ albumin
  • ↑ α1 and α2 globulins
38
Q

Acute Inflammatory response picture

A
39
Q

Chronic inflammatory response

A
40
Q

Liver damage

A
  • Cirrhosis can be caused by
    chronic alcohol abuse or viral
    hepatitis
41
Q

Liver damage affects what values

A
  • ↓ albumin
  • ↓ α1, α2 and β globulins
  • ↑ Ig A in γ-fraction
42
Q

Liver damage picture

A
43
Q

Nephrotic syndrome

loss of what

retention of what

A
  • the kidney damage illustrates the long
    term loss of lower molecular weight
    proteins
    (↓ albumin and IgG – they are filtered
    in kidney)
  • retention of higher molecular weight
    proteins (↑↑ α2-macroglobulin and ↑β-
    globulin
44
Q

Nephrotic syndrome picture

A
45
Q

Monoclonal Gammopathy causes

Production of blank is associated with

Blank can be found in a different what and positions

A

§ Monoclonal gammapathy is caused by
monoclonal proliferation of β-lymphocytal
clones. These “altered” β-cells produce an
abnormal immunoglobulin paraprotein.
§ Production of paraprotein is associated
with benign monoclonal gammopathy
(leucemia) and multiple myeloma.
§ Paraproteins can be found in a different
position: between α-2 and γ-fraction.

46
Q

Monoclonal Gammopathy pictures

A
47
Q

Urine proteins

normal physiology

They appear in blank because

A
  • Proteins found in the urine are from the blood; however, urinary
    proteins can originate from the kidney and urinary tract and from
    extraneous sources such as the vagina or prostate.
  • They appear in the urine because they have passed through the renal
    glomerulus and have not been reabsorbed by the renal tubule
48
Q

CSF proteins

Changes are usually do to what

What should accompany it

mechanism

A

About 95% of the proteins found in CSF result from active
transport of plasma proteins at the blood-CSF barrier.
§ Changes are usually due to lesions, trauma, vascular damage or
infection.
§ CSF protein electrophoresis should also accompany serum electrophoresis

49
Q

Other Applications of electrophoresis used in special chem

A

§ Isoenzyme Electrophoresis
§ CK, LD, ALP & others
§ Lipid Electrophoresis
§ Hemoglobin Electrophoresis
§ Glycoprotein Electrophoresis
§ Nucleic Acid Electrophoresis

50
Q

Hemoglobin and Heme derivatives

Migration is from

In the Blank, Hb is

In blank, Hb is

A

§ Migration is from
cathode to anode.
§ In the Cellulose
Acetate (pH 8.4), Hb is
negatively charged.
§ In Citrate Agar (pH
6.0), Hb is positively
charged.

51
Q

Cellulose acetate and Citrate agar picture

A
52
Q

Immunoelectrophoresis

A

§ Antigen is added to the well and electrophoresis
is carried out. Under the electric field, the
charged antigens get separated based on size
and charge.
§ Following the separation of the antigens, the
antisera is added to the trough and incubated
overnight. During that time the antigen-antibody
reaction causes a precipitation where the
equivalence point is met

53
Q

Rocket electrophoresis

A

§ Rocket Electrophoresis
§ Electroimmunodiffusion
§ Type of immunoassay wherein a single concentration
gradient is established for the antigen, and a voltage
is applied, which drives the antigen from the
application well into a homogeneous suspension of
antibody in the gel.
§ The height of the rocket shaped precipitin line is
proportional to the antigen concentration.

54
Q

Electrophoresis techniques

The tests specimen may be

The primary use of IFE is

A

§ The tests specimen may
be serum, urine, CSF, or
other body fluids
§ The primary use of IFE in
clinical laboratories is for
the characterization of
monoclonal Igs

Lab ops (2 decks)

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