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2.2 clinical chemistry > Plasma Proteins - Introduction and Methods of Analysis > Flashcards

Plasma Proteins - Introduction and Methods of Analysis Flashcards

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1
Q

What percentage of whole blood is plasma

A

55%

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2
Q

What is plasma composed of?

A

Water (92%)

Proteins (4%)

Lipids (3%)

Other solutes (1%)

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3
Q

What proteins are found in plasma
(4)

A

60% albumin

35% globulins

4% fibrinogen

1% regulatory hormones

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4
Q

What does albumin do
(3)

A

Contributes to plasma osmotic pressure

Binds to hormones and inactivates them

Transports lipids

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5
Q

What do globulins do?

A

Transport ions, hormones, immune function

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6
Q

What does fibrinogen do

A

Essential component of clotting system

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7
Q

What are regulatory proteins

A

Enzymes and hormones

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8
Q

How many plasma proteins are there

A

150+

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9
Q

What is the range of sizes in plasma proteins

A

Between 21 kDa and 971 KDa

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10
Q

What is plasma vs serum

A

Plasma = anticoagulant added - contains all clotting proteins

Serum = allowed to clot - clotting removes many proteins especially fibrinogen so the total protein concentration will be lower

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11
Q

What is the role of plasma proteins
(4)

A

They are trapped in the vascular system -> cant fit in capillaries

They provide a colloidal osmotic pressure

They maintain normal blood volume

Maintain normal water load in interstitial fluid and tissues

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12
Q

What is the intracellular compartment

A

Consists of the fluid inside of cells

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13
Q

What is the extracellular compartment

A

Consists of the interstitial fluid and intravascular fluid (plasma)

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14
Q

How is total body water distributed

A

2/3 is in the intracellular fluid compartment
1/3 in the extracellular fluid compartment

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15
Q

What makes up extracellular fluid

A

75% interstitial fluid
25% intravascular fluid/plasma

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16
Q

What proteins are responsible for transport/binding

A

Albumin
Apolipoprotein
Haptoglobin
Thyroxine binding protein

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17
Q

What protein is responsible for oncotic pressure

A

Albumin

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18
Q

List some enzyme inhibitor proteins
(3)

A

Alpha-1-antitrypsin

Cystatin C

C 1-esterase inhibitor

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19
Q

List three immune defence proteins

A

Immunoglobulins

Complement

C-reactive protein

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20
Q

List three acute phase proteins

A

C-reactive protein

Alpha 1 acid-glycoprotein

Serum amyloid A (SAA)

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21
Q

List some proteins with enzymatic activity
(5)

A

Complement C3

Complement C4

Haptoglobin

Cerulopasmin (CER)

Alpha-1-antitrypsin

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22
Q

List some tissue derived proteins
(5)

A

Fibrinogen

Antithrombin

Coagulation factors

Proteins of the fibrinolysis process

Fibronectin

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23
Q

Where are most proteins synthesised

A

The liver

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24
Q

Where are gamma globulins synthesised

A

Synthesised and secreted by plasma cells

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25
Q

Where are plasma proteins synthesised
(3)

A

By the liver cells

By endothelial cells

By blood cells e.g. lymphocytes and plasma cells (immunoglobulins)

26
Q

What influences protein synthesis
(4)

A

Nutritional status

Various feedback mechanisms

Hormonal factors

Genetic factors

27
Q

How much albumin do you make a day

A

About 10g of albumin

28
Q

Where does degradation of proteins occur
(2)

A

Mainly in the liver

Some proteins e.g. albumin are eliminated via the renal system

29
Q

How are proteins broken down in the liver
(3)

A

Proteins are endocytosed by hepatocytes

Plasma proteins are then deglycosylated and then cleaved into amino acids by proteinases and peptidases

This occurs in the lysosomes and cytosol of the cell

30
Q

Where in hepatocytes does protein breakdown occur

A

Lysosomes and the cytosol

31
Q

Describe protein homeostasis
(2)

A

Synthesis of protein must be equal to protein catabolism

i.e. protein production must equal protein loss

32
Q

What affects protein concentrations in blood
(2)

A

Abnormalities -> which arise from a wide variety of unrelated conditions

Alterations in hydration which affect the concentration of proteins but not the absolute amount

33
Q

List the most important groups of proteins
(11)

A

Enzymes

Polypeptide hormones

Antibodies and complement components

Plasma proteins

Apolipoproteins

Haemoglobin

Protein coagulation factors

Structural proteins

Contractile proteins

Storage proteins

Chromosomal proteins

34
Q

What is the main role of polypeptide hormones

A

They regulate metabolism

35
Q

What is the main role of plasma proteins

A

Maintain oncotic pressure

36
Q

Name three structural proteins

A

Keratin

Collagen

Elastin

37
Q

Name a contractile protein

A

Myosin

38
Q

Name a storage protein

A

Ferritin

39
Q

Name a chromosomal protein

A

Basic histones

40
Q

What is the equation for total plasma proteins

A

Albumin + globulins + fibrinogen

41
Q

What is the equation for globulins

A

Total proteins-albumin = globulin level

42
Q

Why do we measure total protein and globulins

A

They give us a general index of overall health and nutrition

The globulins also contain antibodies

43
Q

What is the albumin globulin ration and why do we measure it
(2)

A

Serum albumin/ serum globulin g/l

This ratio can help us identify disorders of serum proteins

44
Q

What is the optimal albumin/globulin ratio

A

1.5 -> 2.2

45
Q

List the 6 methods used to analyse total proteins

A

Kjedahl method

Biuret reaction

Dye binding reactions

UV spectrophotometry

Refractometry

Turbidometric methods

46
Q

What is the Kjedahl Method
(5)

A

This measures nitrogen

We assume that protein contains 16% nitrogen

Any nitrogen containing compounds (proteins) in plasma are digested with sulphuric acid, a catalyst and a salt

Ammonia is produced which is then distilled into a boric acid solution and titrated with acid

Must then correct for non-protein nitrogen by analysing a protein free filtrate as well

47
Q

What is the Biuret Reaction
(2)

A

Most common method used to analyse protein

It is an automated process in labs

48
Q

What is the principle behind the Biuret reaction
(3)

A

Copper reacts in an alkaline solution with peptide linkages to produce a violet coloured complex

This can be measured used a spectrophotometer at absorbance 540 nm

This detects all different types of proteins and is accurate for the range of 10-100g/l

49
Q

What are the downfalls of the Biuret Reaction
(2)

A

Need at least 2 peptide linkages -> will not detect free amino acids

Not sensitive enough for low concentrations i.e. can’t be used for CSF, urine and many body cavity effusions

50
Q

What dye binding methods are used to analyse proteins
(3)

A

Proteins can bind certain dyes resulting in a change of absorption spectrum

Coomassie blue and pyrogallol red are typically used

Different proteins bind different amounts of due therefore the colour change depends on the composition of the protein

51
Q

What are the downfalls of dye binding methods to analyse proteins

A

Difficult to standardise for complex protein mixtures as each protein absorbs a different amount of dye

Some dyes are specific e.g. bromocresol green for albumin

52
Q

What dye is specifically used for albumin

A

Bromocresol green

53
Q

How can UV spectrophotometry be used to analyse proteins
(4)

A

Aromatic amino acids such as phenylalanine and tyrosine absorb light at 280nm

Therefore abs 280 can be used to estimate protein composition e.g. 1g/L Albumin = Abs280 of 0.530

Therefore if your sample reads at 0.530 it contains 1g/L albumin

By knowing the Molar absorptivity of a particular protein its concentration can be determined using Beers Law (A = ebc)

54
Q

What are the downfalls of UV spectrophotometry to analyse protein
(2)

A

Difficult to standardise for complex protein mixtures

Only used for specific proteins such as albumin or IgG

55
Q

Give two examples of aromatic amino acids

A

Phenylalanine and tyrosine

56
Q

What is refractometry used for in the analysis of proteins

A

Used for estimating plasma protein (including fibrinogen) in EDTA plasma

57
Q

What is the principle of refractometry
(2)

A

Measure the refractive index of a sample relative to the refractive index of water

The reading is a measurement of total solids and is only an estimate of protein concentration since there is variation in other serum components e.g. sodium, phosphate, glucose etc which can affect the refractive index as well

58
Q

What is the main downfall of refractometry

A

Lipaemia and moderate to severe haemolysis renders the results invalid

59
Q

What is the turbidometric method of analysing protein
(2)

A

A sensitive method of quantifying protein in low-protein fluids such as CSF or urine

Used when protein level is too low for biuret or refractometer method

60
Q

What is the principle behind turbidometry

A

The process of measuring the loss of intensity of transmitted light due to the scattering effect of particles suspended in a sample

61
Q

How sensitive is turbidometry

A

Sensitive to as little as 60 mg/L of protein

2.2 clinical chemistry (28 decks)

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