Electrophoresis Flashcards

0
Q

What kind of technique is it?

A

Separation

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1
Q

What is it?

A

Movement of charged particles or ions within an electric field

Ions that differ in charge and/or mass will move at different rates.

If they have a common starting point some molecules will move faster than others.

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2
Q

How does it work?

A

If a voltage is applied between two electrodes an electric field (potential gradient) is produced.

E=voltage/ distance between electrodes =V/d (volts/cm)

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3
Q

Particle movement

A

Positive charged ions move to NEGATIVE (cathode) electrode (known as cations) (+)

Negative charged ions move to POSITIVE electrode (anode) known as anions

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4
Q

Movement of the ion is termed…

A

Electrical mobility

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5
Q

Used to seperate..

A

Amino acids, peptides, glycoproteins proteins, nucleotides and nucleic acids and sugars

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6
Q

Factors affecting mobility

A

Size and shape of the molecule and charge
Buffer pH
Medium

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7
Q

How is mobility affected by charge…

A

Net charge of ion will determine which direction it will move

And how much friction from the medium it encounters

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8
Q

How is mobility affected by molecule shape?

A

Structural differences

Globular will migrate differently to fibrous proteins

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9
Q

The supporting medium should be..

A

Inert but can show molecular sieving, absorption and electro osmosis

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10
Q

Examples of medium

A

Paper, acetate, agarose gel, polyacrylmide gel

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11
Q

What is molecular sieving?

A

Associated with gels that consist of pores

Pores restrict flow of ions esp large ones

Smaller ones do not experience resistence and travel further

Separation by molecular size

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12
Q

What is adsorbtion?

A

Rocket shape appearance on medium surface

Caused by retention of molecules and slows down migration of sample

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13
Q

What is electro osmosis?

A

Occurs in media that possesses negative charged groups on surface

Groups attract cations (-) present in buffer

Cations are hydrated

When electric field is applied cations and water are attracted to cathode opposite to flow to anode

Weakly charged ions carried to cathode too

Forms basis of capillarity electro phoresis

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14
Q

Reason for staining

A

Most proteins are colourless

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15
Q

Coloured dyes are used for

A

Paper, cellulose acetate or gels

16
Q

Fluorescent dyes are used for…

A

Ethidium bromide - DNA gels

17
Q

Example of scanning densitometry usages

A

Multiple myeloma- determine presence of abnormal serum proteins and haemolytic variants