Clinical Chemistry I - Laboratory Analysis of Protein (Module 1) Review

Hypoproteinemia

Total plasma protein level less than reference interval.
Occurs in any condition where a negative nitrogen balance exists.

Hyperproteinemia

An increase of total plasma proteins above reference interval.

Hypoproteinemia

Due to:
◦Excessive loss (renal disease, fluid loss, burns)
◦Decreased intake (nutritional)
◦Decreased synthesis (liver)
◦Accelerated breakdown (trauma)
◦Immunodeficiency (decreased synthesis)

Hyperproteinemia

Due to:
◦Dehydration - Relative change when concentration of proteins is elevated due to decreased volume of water
◦Excess production of proteins (g-globulins) - Multiple myeloma, Waldenstrom's macroglobulinemia

Total Protein testing

Very common, routine lab test utilizing serum or plasma; reflects nutritional status, hepatic and renal function, overall protein value; values low at birth, reach adult levels by age 3

6.5-8.3 g/dL

Total protein (TP) reference range/interval

Total protein measurement methods

Biuret, dye binding, turbidimetry and nephelometry

Biuret method

Method of total protein measurement based on cupric ions (Cu2+) chelating with peptide bonds - Forms violet colored complex where Beer's law can be used

Dye binding method

Method of total protein measurement based on the affinity of protein with a specific dye.
- Bromocresol green (BCG) - can be falsely elevated by Hb and alpha globulins
- Bromocresol purple (BCP) - Preferred, less interferents
- Amido black

Turbidimetry and nephelometry method

Method of measuring total protein based on light transmission or light scatter due to particles present

3.5-5.5 g/dL

Albumin reference range/interval

Albumin/Globulin ratio

Total protein minus (-) albumin = globulins,
Albumin divided (/) by globulins =

1.1-1.8 g/dL

Serum A/G ratio reference range/interval

Low A/G Ratio

Indicative of autoimmune disorder, cirrhosis, kidney disease

High A/G Ratio

Indicative of leukemia, liver disease, hyperthyroidism

Serum Protein Electrophoresis (SPE)

- Proteins at pH 8.6 carry a net negative charge and migrate toward the positive terminal, the anode
- After separation, fix and stain the proteins
- Migration depends on size, shape, temperature, pH, and ionic strength of the test system
Overall net negative charge on the protein at pH 8.6

High resolution electrophoresis

Electrophoresis that uses higher voltage and a cooling system to increase separation into 15 bands

Capillary electrophoreesis

Separation of molecules takes place in buffer filled silica capillary tubes

Isoelectric focusing

Zone electrophoresis that separates on basis of pI (isoelectric point)

Immunofixation electrophoresis

Electrophoresis specific for monoclonal gammopathy isolation on gel

CSF proteins

95% ____ _________ is from plasma active transport across blood-brain barrier
Ratio of serum albumin with CSF albumin identifies the degree of permeability

CSF proteins

Elevated based on 2 factors:
- Increased permeability of blood-brain barrier -> Multiple sclerosis, bacterial viral & fungal meningitis, neoplasm/tumor
- Synthesis by inflammatory cells (neurosyphilis, hematoma, heemorrhage)

CSF protein electrophoresis

Used in detecting multiple sclerosis - unexpected oligoclonal bands in gamma-globulin region

Alpha 1 Globulins

Alpha-1 Antitrypsin (AAT),
Alpha-1 Fetoprotein (AFP), Alpha 1 Lipoprotein

Alpha 2 Globulins

Haptoglobin, Ceruloplasmin,
Alpha-2 Macroglobulin

Beta Globulins

Transferrin, hemopexin, Beta-2 Microglobulin (B2M), C3, C4, Beta Lipoproteins, C-Reactive Protein (CRP), Fibrinogen (Plasma only)

Gamma Globulins

IgG, IgM, IgE, IgA, IgD

Transport binding

Function for albumin, prealbumin, haptoglobin, hemopexin, transferrin

Oncotic pressure

Function for albumin

Immune defense

Function for Immunoglobulins, complement, CRP

Acute phase reactants

C-Reactive protein (CRP), Alpha 1 Antitrypsin (AAT), ceruloplasmin - markers of inflammation

Nephrotic syndrome

- Loss of albumin
- Increased alpha 2 macroglobulin in α2 region since it is too large to be excreted, is retained
- Takeaway: Decrease of all fractions with the exception of α2

Acute Inflammation

Increase in acute phase proteins in α2 region
Takeaway: Decreased Albumin, increased α1 & α2, normal gamma

Inflammation

Alpha 1 or Alpha 2 globulin fraction increase means:

Chronic inflammation

Decreased Albumin, increased alpha 2 (α2), increased Gamma

Hypoglobulinemia

No Gamma (ɣ) globulin peak

Hyperglobulinemia (monoclonal gammopathy)

Spike at Gamma (ɣ) globulin peak

Hepatic Cirrhosis

Decreased albumin production
Classic beta-gamma bridge due to immunoglobulin subclasses

Alpha1 Antitrypsin Deficiency

Decreased Alpha 1

Severe Hepatic Disease

Decrease in all zones except gamma (increased) due to a failure of the liver to synthesize proteins (Decreased albumin, alpha 1 and 2, beta, Increased gamma)