Biochemical Profiling: Protein Flashcards by Adriane Gorski

What tube is used to sample protein on the CBC? Chem? What sample is used?

CBC - purple top or green top: plasma

Chem - red top: serum

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What is different about the samples used for calculating protein on the CBC vs. Chemistry?

CBC – has all clotting factors (fibrinogen, Factor V and VIII)
Chemistry – no clotting factors –> lower protein value

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Which result will tend to have a higher TP, CBC or Chem?

CBC TP

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What do you use for a STAT chem?

green top (heparin plasma)

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What protein is calculated?

Globulins = TP - Albumin

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What are 3 interferences with refractometry ?

lipids
glucose, BUN, NaCl
hemolysis

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What are some interferences with chemistry methods? What is affected more?

Lipemia
hemolysis
possibly icterus

TP tends to be more affected than albumin

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What are some physiological influences on protein values?

Age of animal
Nutritional status
Pregnancy --> lactation
Hormonal 
Gender (minimal)

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What happens with TP in young animals? Old?

Young: low end of ref range or below, increase gradually for ~ 6 months

Old: trend of increasing TP (decreased Alb, increased globulins)

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What are albumin levels in young animals than adults? When do globulin values jump in young animals?

0.5 - 1.0 g/dl lower than adult

weaning and vaccination

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What are 3 possible mechanisms for pregnancy or lactation causing decreased protein levels?

dilutional effect of increased blood volume (pregnancy)
Decreased synthesis?
Increased catabolism during high energy needs?

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What hormones are anabolic to proteins? catabolic? What is the exception?

Anabolic: testosterone, estrogen, growth hormone
Catabolic: thyroxin, cortisol**

exogenous glucocorticoids may cause upward shifts in albumin

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What is fibrinogen produced by? Why is it used as an indicator of inflammation?

liver

acute phase inflammatory reactant

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Does fibrinogen increase or decrease in response to inflammation? Why?

Increase

acute phase inflammatory reactant

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What does fibrinogen do in response to DIC, increase or decrease?

Decrease

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What are some differentials for increased production of fibrinogen (inflammation)? Non- inflammatory?

INFLAMMATORY

Infectious
Traumatic
Neoplastic

NON-INFLAMMATORY
1. Dehydration

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What are 5 mechanisms for hypofibrinogen?

DIC (increase utilization)
Hepatic failure (decrease production)
Protein-losing disease (losses, concrrent with other proteins)
Congenital (rare)
Clotted blood! (EDTA/citrated samples with clots)

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What is the purpose of the total plasma protein: fibrinogen ratio?

Help determine whether fibrinogen values reflect relative shifts (with other proteins, dehydration) or whether there is a specific increased production of fibrinogen

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What are 2 interpretive dilemmas for evaluating fibrinogen alone?

In Hypoproteinemias (TP): may be underestimated because of general protein losses such as in hemorrhage or protein losing diseases
In Hyperproteinemias (TP): values may be overestimated because of overall relative increase in proteins due to dehydration.

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How do you calculate TPP/Fibrinogen ratio? What is a very important part of this formula?

TPP (g/dl) - Fibrinogen (g/dl) / Fibrinogen (g/dl)

TPP = total PLASMA protein

IMPORTANT
everything must be in g/dl. Convert fibrinogen values from mg/dl to g/dl
700mg/dl = .70 g/dl

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How do you interpret the TPP/Fibrinogen ratio?

< 10 Active inflammation
10-15 Gray zone (ambiguous)
> 15 Normal fibrinogen (changes “relative”, associated with fluid alterations)

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What does an ambiguous TPP/fibrinogen ratio tell us?

We can’t say whether it is relative increase or from inflammation. We need to look at other modalities to say it is inflammation (leukon)

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Where could dysproteinemia’s mechanistic localization be?

Production
Loss
Third space
Relative change (fluid related shifts)

What can be some causes for dysproteinemia loss?

Hemorrhage
GI tract
Urinary tract
Skin (burns)

What are some production causes that may result in dysproteinemia? More looking for the site of production....

Liver Lymphoid organs Mononuclear phagocyte system (macrophages) - minor contribution

What are two relative changes that may cause dysproteinemia?

Dehydration | Hemodilution

What is the differential list for dysproteinemic state?

Increased protein Decreased protein Mixed protein pattern (hypoalbuminemia + hyperglobulinemia)

What are three causes of hyperalbuminemia (selectively)?

1. relative increases - dehydration (major cause) 2. glucocorticoids (possible increase) 3. Laboratory interferences with the BCG method (hemolysis, lipemia, +/- icterus)

What are causes of hyperglobulinemia (selectively)?

1. Increased synthesis - chronic antigenic stimulation (hepatic insufficiency, fungal diseases, FIP) --> immunoglobulin production - Active inflammation --> mild increase only (acute phase proteins) - Lymphoproliferative dz: immunoglobulin production

Cause for Panhyperproteinemia?

Dehydration

Causes for Hypoalbuminemia

1. Decreased synthesis - active inflammation - hepatic insufficiency** - malnutrition, malabsorption (both, only in severe cases) 2. Increased loss - Protein losing glomerulopathy** - early/mild PLE 3. Sequestration/Redistribution - Exudation

Causes for hypoglobulinemia

1. Decreased synthesis - Immunodeficiency --> congenital or acquired (FPT) - Liver: decreased functional mass

What are causes of panhypoproteinemia?

1. Hemodilution 2. increased loss - acute blood loss - PLE** - severe skin lesions (burns, exudation) - +/- protein losing glomerulopathy ** 3. Sequestration - body cavity effusions (dilutional)

What does SPE stand for?

Serum Protein Electrophoresis

What is SPE?

The separation of charged protein particles in an electric field

Does SPE separation rely more on charge or size for migration pattern relative to other proteins?

Charge

What are the 5 fractions that proteins are usually grouped into for SPE?

1. Albumin 2. Alpha-1 3. Alpha-2 4. Beta 5. Gamma

What gamma proteins does the SPE measure?

IgG, IgA, IgM

What beta proteins does the SPE measure?

``` C-reactive protein serum amyloid A transferrin C3, C4 IgA, IgM Fibrinogen in plasma samples only ```

What SPE fractions measure both IgA or IgM? Fibrinogen? IgG?

Beta and Gamma -- IgA and IgM Beta - fibrinogen Gamma - IgG

What two negative acute phase proteins does the SPE look at?

Transferrin | Albumin

When do you do an SPE?

When you are not satisfies with the protein measurements found on CBC or chem. Those 2 assays are very poor at being specific.

What are the 4 indications for doing SPE?

1. Help differentiate causes of hyperproteinemia 2. Help characterize unexplained hypoprtoeinemia 3. Monitoring dz activity/progression 4. More accurate measurement of albumin when globulins cross-react on BCG assay

Name the 3 parts you should get when asking for a SPE?

1. Bands separated on gel 2. Electrophoretogram 3. Associated data

What two things do you need to look at when interpreting an SPE?

1. electrophoretogram pattern | 2. quantitative data

What does a large peak mean?

A peak may be large and still represent a decreased amount of protein in hypoproteinemic states

What are three key patterns when looking at the SPE?

1. Active inflammation 2. Monoclonal gammopathy (regardless of whether the spike is in beta or gamma region) 3. Polyclonal gammopathy

What does an active inflammation look like on the SPE electrophoretogram?

alpha protein spike or an increase in total alpha proteins | occasionally have increased beta protein or beta spike

When there is a monoclonal gammopathy what should you think of first?

lymphoid neoplasia

What does a monoclonal gammopathy look like on the SPE electrophoretogram? What does it generally represent? What type of disease is it most commonly due to?

narrow-based spike (< albumin width) in beta or gamma region generally represents a single class of Ig -- single clone of cells causing its increase lymphoproliferative disease (multiple myeloma)

When you see a polyclonal gammopathy what should you think of first?

Chronic antigenic stimulation/ inflammation

What does a polyclonal gammopathy look like on the SPE electrophoretogram?

Broad based increase (>1.5x albumin width) in beta gamma regions Beta gamma bridging (no clear separation b/w the two regions)

What chronic antigen stimulations is polyclonal gammopathy typically associated with?

FIP Ehrlichiosis chronic pyelonephritis possibly systemic fungal disease

What is beta gamma bridging suggestive of?

hepatic insufficiency

How do you calculate the protein amount from the SPE data?

Area under peak of electrophoretogram x TP (chem) = true protein amount

What other protein can you expect to see changes in with a polyclonal gammopathy?

hypoalbuminemia