IgG IEF

Diagnosing Multiple Sclerosis

Multiple Sclerosis

• CNS disorder

• Scar formation on outside of nerve cells of brain and spinal cord

• Inflammation destroys covering of nerve cells leaving scar tissue

• Nerve cells cannot transmit impulses

Prevalence

• Onset at age 15 to 50 years

• 70% female

• Temperate climate predominant

Symptoms

• Optic neuritis• Numbness / weakness in

extremities• Instability in walking• Tremors• Loss of bladder control• Heat intolerance• Fatigue

Diagnosis by Exclusion

• Encephalitis• CNS Vasculitis• Lyme Disease• Behet Syndrome• Sarcoidosis• Syphilis• Leukodystrophies

• Lupus Erythematosus• Spastic Paraparesis• Vitamin B-12 Deficiency• Sjogren’s Syndrome• Hereditary Degenerative

Disorder• Multifocal

Leukoencephalopathy

Criteria for Diagnosis

History of two attacks with

positive oligoclonal bands

or

increased IgG in CSF

Criteria for Diagnosis

History of two attacks with

no laboratory abnormality

Criteria for Diagnosis

History of two attacks with

clinical and paraclinical evidence of lesions, oligoclonal bands, and

increased IgG in CSF

Criteria for Diagnosis

History of two attacks with

clinical evidence of at least one lesion, and clinical or paraclinical

evidence of another lesion

IEF Theory

• Stable pH gradient with carrier ampholytes in an electrical field

IEF Theory

• Stable pH gradient with carrier ampholytes in an electrical field

• Proteins exhibit net + or – charge

IEF Theory

• Stable pH gradient with carrier ampholytes in an electrical field

• Proteins exhibit net + or – charge

• Migrate toward electrode of opposite charge

IEF Theory

• Stable pH gradient with carrier ampholytes in an electrical field

• Proteins exhibit net + or – charge

• Migrate toward electrode of opposite charge

• Rate of migration ↓ as it reaches isoelectric point

IEF Theory

• Stable pH gradient with carrier ampholytes in an electrical field

• Proteins exhibit net + or – charge

• Migrate toward electrode of opposite charge

• Rate of migration ↓ as it reaches isoelectric point (pI)

• Protein focuses at pI

IEF Theory• Stable pH gradient with carrier

ampholytes in an electrical field• Proteins exhibit net + or – charge• Migrate toward electrode of

opposite charge• Rate of migration ↓ as it reaches

isoelectric point (pI)• Protein focuses at pI; some drift

but regains charge and migrates back

IEF Procedure

Step 1: Focus proteins

IEF Procedure

Step 2: Transfer proteins to nitrocellulose membrane

IEF Procedure

Step 3: React with IgG antisera

IEF Procedure

Step 4: React with chromagen Step 5: Air dry

Total Time: 2.5 to 3.0 hours

Interpretation

Compare CSF and Serum

Looking for intrathecal synthesis

indicated by

oligoclonal bands in CSF

but not in serum

Interpretation

Negative – no oligoclonal bands in CSF

Interpretation

Mirror Image – same bands in CSF and serum

Interpretation

Positive – Oligoclonal bands in CSF but not in serum

Interpretation

Positive oligoclonal banding is not diagnosis of MS