1
Q
General concepts
A
> 1,000 characterized in the serum
- plasma: albumin, globulins (fibrinogen, clotting factors)
2
Q
Serum does not contain ______
A
Fibrinogen
3
Q
Most of the plasma proteins are synthesized by ________
A
Hepatocytes
- exceptions: Ig (B cells and plasma cells)
- proteins contribute to colloid oncotic pressure
4
Q
Protein dyscrasia
A
Presence of abnormal protein
- abnormal structure
5
Q
Dysproteinemia
A
Presence of normal protein at abnormal concentration
- selective or nonselective dysproteinemia
6
Q
Nonselective hyperproteinemia
A
All protein concentrations are increased at the same rate
7
Q
Selective hyperproteinemia
A
Total protein concentration is increased and some protein concentration are increased more than others
8
Q
Nonselective hypoproteinemia
A
Total protein concentration is decreased and all proteins are decreased
9
Q
Selective hypoproteinemia
A
Total protein concentration is decreased and some protein concentration are decreased more than others
10
Q
Selective electrophoresis
A
To determine if the condition is selective or nonselective
- albumin and globulins decreased or increased at the same rate (nonselective)
- albumin and globulins decreaed or increased but not proportional (selective)
11
Q
Other than dehydration, the most common cause of dysproteinemia is _____
A
Inflammation
12
Q
Positive acute phase proteins
A
- SSA and CRP are increased in less than a day
- major APPs: fibrinogen, CRP, SAA, haptoglobin, a1 acid glycoprotein, ferritin, ceruloplasmin
13
Q
Negative APP
A
Decrease plasma or serum concentration due to inflammatory process
- major negative APPs: albumin, transferrin
14
Q
Delayed response proteins
A
Serum or plasma concentration increases 1-3 weeks after onset of inflammation
- major delayed response proteins: Ig, complement factors (C3)
15
Q
Refractometer
A
Degree of water refraction in a solution is proportional to the quantity of solids
- most of the solids in plasma are proteins
16
Q
Refractometer - interferences
A
- other substances (urea, glucose, Na, Cl)
- lipemia increases refractive index
- hemolysis does not interfere
- bilirubin does not interfere
17
Q
Biuret reaction measures ______
A
Total protein
- colorimetric: amount of color change is proportional to the amount of protein
- interferences: hemolysis (positive), dextran (positive)
18
Q
Albumin concentration
A
BCG dye binding reaction
- colorimetric
- interferences: binding to Ig when albumin concentration is low, hemoglobin (positive), triglycerides (negative), heparin (positive)
19
Q
Globulin concentration
A
Determined by subtraction
- total protein - albumin = globulins
- is the concentration of all other proteins other than albumin
20
Q
Serum protein electrophoresis (SPE)
A
Migration through a acetate cellulose or agarose gel
- most proteins migrate towards the anode –> albumin is the smallest and negatively charged and migrates further, some Igs are positively charged and migrates towards the catode
- separate in 4-6 groups of one or more band
21
Q
Albumin is a negative acute phase protein because it _______
A
Decreases concentration in inflammation
22
Q
SPE - species variations
A
- 1 mg/dL is the minimum detectable concentration
- most dogs, cats, and horses: a1, a2, b1, b2, and y
- cattle: a, b and y
- concentration of proteins are calculated after the stained gel is scanned with a densitometer –> area under curve is the total stained protein
23
Q
Concentration of each group is its specific percentage times _______
A
The total protein concentration (measured using biuret technique)
24
Q
Diagnostic value
A
- differentiating into selective or nonselective
- scanning for a monoclonal gammopathy
- helping to assess albumin concentration when globulins interfere with BCG
25
________ is the most common cause of hyperproteinemia
Hemoconcentration
- caused by loss of plasma water
- vomiting, diarrhea, impaired renal concentration, sweating, increased vascular permeability
- if TP 7 g/dL, then in a 10% dehydrated animal it would be 7.7 g/dL
- nonselective
- other: erythrocytes, prerenal azotemia, hyperstenuria
26
Increased protein synthesis
Inflammation is a common cause of hyperproteinemia
- does not always cause hyperproteinemia
- increase production of positive APP and delayed response proteins (mostly by hepatocytes)
- increased production of positive APP: within hours and persists until inflammation is resolved
- increased plasma conc in 2 days
- increase of [fibrinogen] and [haptoglobin] can cause hyperproteinemia
27
Hyperproteinemia due to inflammation
Decreased production of negative APP
- albumin half life: 8 days in dogs, 19 days in horses
- hypoalbuminemia due to inflammation: several days in dogs, 2 weeks in horses
- transferrin: half life not well established, decreased conc after at least a week
28
Inflammation leads to increased production of ________
Delayed response proteins
- 1-3 weeks after the onset of inflammation
- Ig, mostly IgG and complement
- C3 usually <1.0 mg/dL
- IgG mild: <1.0 mg/dL to marked >4.0 mg/dL
29
Other findings with hyperproteinemia due to inflammation
- anemia of inflammatory disease
- neutrophilia, neutropenia
- lymphocytosis, lymphopenia
- monocytosis
30
B lymphocyte neoplasia
B cells may produce Ig
- plasma cell neoplasia: multiple myeloma (most frequent cause) or extramedullary plasmacytoma
- lymphocyte neoplasia: lymphoma or lymphocytic leukemia
31
Expected dysproteinemia pattern with B cell neoplasia
- IgG migrates with g globulin fraction
- IgM or IgA with B-y or B globulin fraction
- atypical pattern seen due to protein degradation or complex with other protein, Ig complexes, and incomplete Ig
- other [Ig] are decreased
- mild-moderate hypoalbuminemia due to decreased production or neg feedback by oncotic pressure receptors in hepatocytes
32
B cell neoplasia - associated findings
- Bence Jones proteinuria: light chains of Ig in the urine
- hyperviscosity syndrome: high Ig leads to increase plasmatic viscosity
- hypercalcemia
- AL amyloidosis caused by Ig light chains
- PCR for Ig variable region gene rearrangement can be used for diagnosis
33
Blood loss
Increased loss from vascular space, hypoproteinemia occurs when remaining blood is diluted
- extracellular extravascular fluid shifts to intravascular
- panhypoproteinemia (nonselective)
- anemia (depends on severeity and duration of hemorrhage)
34
Protein losing nephropathy
Immune complex or amyloid deposition in glomeruli --> retraction of podocytes or loss of selective permeability of glomerular basal membrane --> protein loss
- if protein loss > protein production then hypoproteinemia
- larger proteins are not lost, so it is a selective process
- glomerulopathy: glomerulonephritis or amyloidosis may be present
35
Protein losing nephropathy - lab findings
- proteinuria, mostly albuminuria
- moderate-marked hypoproteinemia with moderate-marked hypoalbuminemia
- serum electrophoresis consistent with selective protein loss
- possible renal failure
- possibly hypercholesterolemia if nephrotic syndrome present
36
Protein losing enteropathy
Protein rich intestinal secretions (digestion and resorbed thru lymphatics) --> if reabsorption is impaired (SI mucosal disease, or lymphatics disease) protein lost in feces
- if protein loss > protein production, then hypoproteinemia
- protein exudation and decreased intake
- blood loss, intestinal parasites
37
Protein losing enteropathy - lab findings
- mild-marked hypoproteinemia
| - serum electrophoresis: nonselective
38
Protein losing dermopathy
Thermal or chemical burns --> protein exudation from cutaneous lesions
- if protein loss > protein production, then hypoproteinemia
- after 2 days findings, will include APP shifts
39
Protein losing dermopathy - lab findings
- early: nonselective hypoproteinemia
| - later: nonselective hypoproteinemia masked by acute or chronic inflammatory dysproteinemia
40
Plasma loss (peritonitis, pleuritis, vasculitis)
- pleuritis/peritonitis: inflammation --> protein loss in pleural or peritoneal cavities --> hypovolemia --> shift of fluids from extracellular extravascular to intravascular --> dilution of remaining proteins (hypoproteinemia)
- aka: 3rd space loss
- vasculitis: similar, proteins lost to perivascular interstitial tissue adjacent to affected vessel
41
Plasma loss - lab findings
- early: nonselective hypoproteinemia
| - late: nonselective hypoproteinemia masked by acute or chronic inflammatory dysproteinemia
42
Hepatic insufficiency (hepatic failure)
Marked decrease in hepatic functional mass (<20% remaining) --> decreased production of nearly all plasma proteins (except Ig)
- normal protein catabolism combined with decreased production = hypoproteinemia
- decreaed synthesis and/or increased catabolism
43
Hepatic insufficiency - disorders
- cirrhosis
- hepatic necrosis or inflammation (not acute)
- portosystemic shunt (hepatic atrophy)
- extensive liver damage due to neoplasms
44
Hepatic insufficiency - lab findings
- hypoproteinemia, hypoalbuminemia, normoglobulinemia, hypoglobulinemia
- SE: nonselective, B2 and y may be increased
45
Malabsorption or maldigestion
Deficient intake of carbs, lipids and proteins --> once depleted storages, normal catabolism and use of animo acids for gluconeogenesis --> hypoproteinemia
46
Malabsorption
Small intestine diseases with generalized mucosal involvement
47
Maldigestion
Exorine pancreatic insufficiency (chronic pancreatitis or atrophy) leads to deficiency in protease, lipase, and amylase
48
Malabsorption or maldigestion - lab findings
Hypoproteinemia, hypoalbuminemia, normo or hypoglobulinemia
| - SE: nonselective
49
Cachectic states
Protein catabolism exceeds production --> hypoproteinemia
- before hypoproteinemia --> protein production (mainly albumin) at expenses of body fat and muscle mass
- chronic diseases: infectious and malignant neoplasia
- marked malnutrition or starvation
50
Cachectic state - lab findings
- hypoproteinemia, hypoalbuminemia, normo or hypoglobulinemia
- SE: nonselective
51
Lymphoid hypoplasia/aplasia
B cells produce Ig and not other major plasma proteins
- decreased Ig production in face of normal concentration of other proteins --> mild hypoproteinemia (hypoglobulinemia), normoalbuminemia
- SE: decreased gamma group
- combined immunodeficiency (arabian, appaloasa, basset hounds, corgis, jack russell)
- infectious diseases, chemotherapy
52
Failure of passive transfer
Failure to absorb immunoglobulins from colostrum --> decreased plasma Ig --> hypoproteinemia, hypoglobulinemia
- inflammation can mask the hypoproteinemia
- calves with [IgG] of less than 1,000 mg/dL
- [serum TP] fo 5.0 g/dL or less 83%
53
Hemodilution
Increased ECF is uncommon (by itself) to cause hypoproteinemia
- worsens hypoproteinemia from other causes
- due to: excess IVF, edematous disorders, excessive ADH secretion, use of plasma expanders
54
Hyperalbuminemia due to hemoconcentration
Dehydration, decreased ECF
- decreased water --> increased concentration of all blood contents
- most common cause of hyperalbuminemia
55
Hyperalbuminemia due to induced synthesis by _____
Glucocorticoid therapy
- increase in 4-5 days and 4 weeks (dogs and cats) of treatment
- maybe increased production or increased life span
56
Falsely increased albumin using BCG method
BCG dye binds preferentially to albumin but also to alpha and beta globulins
- heparinized plasma samples can lead to increase due to binding to fibrinogen
- may mask hypoalbuminemia
57
Hypoalbuminemia along with hyperproteinemia
Not due to hemoconcentration
- inflammatory: negative APP, takes 1-2 weeks to appear, concurrent B cell neoplasia due to inflammatory cytokines, increased colloidal osmotic pressure or damaged tissues (liver, kidney, intestines)
58
Hypoalbuminemia along with hypoproteinemia
Decreased production or loss or both
59
Hyperglobulinemia
Frequently with hyperproteinemia but possible with normoproteinemia due to hypoalbuminemia
- commonly caused by hemoconcentration, inflammation (chronic), B cell neoplasia
- may be result of increases in one or several globulins
- SPE: differentiate hemoconcentration from inflammation and B cell neoplasia, patterns of inflammation and B cell neoplasia may overlap
60
Hypoglobulinemia
Other than neonates, commonly with hypoproteinemia and occasionally with normoproteinemia
- decreased production: hepatic insufficiency and lymphoid hypoplasia/aplasia
- loss: blood loss, intestine mucosa, skin
61
Positive acute phase proteins
Any injury that causes inflammation can increase positive APPs
- can increase globulins and TP
- individually, magnitude can vary
62
Why use positive APPs
- insensitivity of other tests (pyrexia, neutrophlia)
| - provide another method of monitoring inflammation for therapeutic decisions and prognostic
63
Positive APPs - factors to consider
- persistence of increase of positive APPs in chronic inflammation
- may be processes decreasing concentration of positive APPs at the same time of inflammation --> disorders that cause loss of plasma proteins, activation of plasmin or thrombin (decreasing fibringogen), intravascular hemolysis
64
Fibrinogen
Plasma protein produced by hepatocytes
- used for production of fibrin by thrombin
- positive APP but consumption may take place at the same time masking increased production
65
Fibrinogen - analytical process
Heat precipitaiton
- measure [TP] in an aliquot of plasma sample --> heat precipitate fibrinogen in a second aliquot and centrifuge the sample to separate fibrinogen --> measure [TP] again --> [fibrinogen]=1st [TP] - 2nd [TP]
- adequate for documenting hyperfibrinogenemia, but not hypofibrinogenamia
- subtracting serum [protein] from plasma [protein] does not work since other solutes are released and/or consumed during coagulation
66
Hyperfibrinogenemia
Due to: hemoconcentration or inflammation
- TP may be increased, but also WRI
- fibrinogen increases are more consistent than TP in inflammation
67
Plasma protein to fibrinogen ratio (PP:F ratio)
Ratio helps to differentiate hyperfibrinogenemia of inflammation from hemoconcentration
- ratio varies among species and age groups
68
Plasma protein to fibrinogen ratio (PP:F ratio) - species variation
- cattle: >15 likely dehydration, <10 likely inflammation
| - horses: >20 likely dehydration, <15 likely inflammation
69
PP:F ratio - additional factors
- concurrent inflammation and dehydration will make interpretation difficult
- dehydration by itself will cause mild increases in [fibrinogen]
- guidelines not adequate for calves and foals
- guidelines not adequate for cattle and horses with pathologic process that causes hypoproteinemia
- accuracy of refractometer is 0.1 mg/dL, so ratio will be more accurate with higher [fibrinogen]
70
Except for fibrinogen, ______ are preferred
Serum samples
| - relative stable to freezing
71
Variety of tests are used depending on APP
- serum C reactive protein
- haptoglobin
- serum amyloid A
- a1: acid glycoprotein
- ceruloplasmin
- ferritin
72
Causes other than inflammation for APP concentration changes
- [CRP] in pregnant bitches
- glucocorticoids can increase [haptoglobin] in dogs
- anti-helmintic drugs can increase [haptoglobin]
- phenobarbitol in dogs may increase
- lactation in cattle increases [CRP]
- increased [ferritin] with increased iron stores
- increased [ferritin] in horses after exercising
73
Reasons to measure Ig
Most common cause to assessing Ig is to determine passive transfer from mares to foals and cows to calves
- congenital or acquired immunodeficiency
- monoclonal gammopathies
74
Failure of passive transfer - process
Placentaiton in horses and calves prevent in utero transfer of Ig
- need to ingest colostrum before gut closure to obtain maternal Ig
- neonate foals have no plasma IgG
- foals and calves have limited ability to produce IgM
- after colostrum intake, [TP] increases due to IgG, but also other APP
- Ig uptake is mediated by Fc receptor on epithelial cells
75
Failure of passive transfer causes
Lack of colostrum intake (too weak to nurse)
- inadequate IgG in the colostrum (low conc or low amount of ingested colostrum)
- failure to absorb (after gut closure) and potentially due to different haplotypes of Fc receptor in calves
76
Establishing FTP - foal guidelines
Foal guidelines: sample from 18 to 48 hrs after birth
- FTP when IgG < 400 mg/dL
- [IgG] >800 mg/dL as adequate is recommended as criterion
- [IgG] <200 mg/dL is complete FTP
- [IgG] <800 mg/dL and >200 mg/dL is considered partial FPT
77
Establishing FTP - calve guidelines
Sample from 1 and 8 days after birth
- [IgG] > 1000-1600 mg/dL as adequate passive transfer
- [IgG] <500-800 mg/dL is complete FTP
- [IgG] <1600 mg/dL and >800 mg/dL is partial FPT
78
Radial immune diffusion (RID)
Most used
- not the most accurate
- takes 24 hrs for results (time for Ig to diffuse thru the gut)
79
Glutaraldehyde coagulation test
- inexpensive
- not reliable with total blood
- for foals good to excellent agreement
- for calves may require dilution of the sample
80
Latex agglutination
Latex beads covered with anti equine IgG agglutinate in the presence of IgG
- semiquantitative test good agreement with RID
- not as good as glutaraldehyde coagulation test
- moderately expensive but takes only 10 minutes
