Laboratory Handling of the Renal BiopsyDr. Issam Francis

Kuwait

4th SSN Annual International Conference, Riyadh, April 2009

The Renal Biopsy Historical backgrounds

• 1934 Percutaneous kidney biopsy (tumors) • 1951 first kidney biopsy for medical disease • 1953 Introduced to US• 1955 first renal clinicopathology working group

Modern Times:• Real-time ultrasound guidance• Transjugular needle biopsy• Spring-loaded biopsy gun• CT-Guided biopsy• EM• IHC

Needle Biopsy Open Biopsy

Open Biopsy

The Renal Biopsy Laboratory handling

EM Paraffin Sections IF

Under a stereomicroscope

The Renal Biopsy Laboratory handling

EM Paraffin Sections & IHC (no IF)

Under a stereomicroscope

The Renal Biopsy Laboratory handling

1- fixation (Immediate):• 10 % NB Formalin (paraffin sections)• 4% Gluteraldehyde (EM)• No fixation (Immunofluorescence)

2- Paraffin sections cut at 3 u thickness

3- Stains: HE PAS GMS TC CR …..HE PAS GMS HE TC HE PAS GMS HETC HE HE PAS GMS HE TC PAS

GMSHE TC (CR, Microbial stains, others.)

4- Immunohistochemistry (IG, C, other antigens)

5- Immunofluorescence (IG, C)

The Renal Biopsy Laboratory handling

• HE• PAS• JNS• TC• HE• PAS• JNS• TC

3-4 microns24 Sections

Extra sectons for CR, Microbial stains, IHC etc.

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The Renal Biopsy Laboratory handling

Biopsy adequacy:– Cortex and medulla– 1-2 glomeruli EM– 3-5 glomeruli IF– 6 glomeruli PS (native kidney)– 10 glomeruli PS (renal allograft)

The Renal Biopsy Manual vs. automated

05

1015202530354045

Man

ual

Au

tom

ated

# of cores

Totalglomeruli

Glomeruliper core

Biopsy Fixation

10% Neutral Buffered Formalin Solution:Why?1. Cheap2. Commonly available3. Suitable for:

– All histological stains– Immunohistological methods (not IF)

4. Reversible: Possible to transfer to another fixative for electron microscopy.

Morphological examination:1. Glomeruli2. Tubules3. Interstitium4. Blood vessels

Renal Biopsy Morphological Examination

Primary Site of Renal Pathology

Glomerulus• Glomerular pathology:

– Inflammation– GBM changes– Scarring– Abnormal deposits– Cellularity

Tubules • Tubular pathology :

– Cellular injury– Regeneration – Atrophy – Casts

Interstitium • Interstitial

pathology:– Cellular infiltrates– Edema/fibrosis

Vascular disease • Vascular pathology:

– Inflammation– Sclerosis– Hyalinosis– Thrombosis

? Stain

Activity of disease:• Cellular proliferation• Crescent formation• Necrotizing lesions• Inflammation

Chronicity of disease :• Tubular atrophy• Fibrosis• Vascular sclerosis

Renal pathology report Disease Stage

The Renal Biopsy Stains1. HE General

2. PAS Basement M. & Mesangial matrix

3. Trichrome Fibrosis

4. Silver Basement M. & Mesangial matrix

5. Congo red Amyloid

6. MSB Fibrin

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Masson’s trichrome

Structure/Component PAS Jones Masson’s trichrome

Basement membrane Red Black Deep blue

Mesangial matrix Red Black Deep blue

Interstitial collagen Negative Negative Pale blue

Cell cytoplasm (normal) Negative (most) Negative Rust/orange

Immune complex Negative Negative Bright red

Amyloid Negative Negative Light blue

Tubular casts Red Gray to black Light blue

Staining characteristics of selected normal and abnormal renal structures

SATINING OF RENAL TISSUE COMPONENTS

FEATURE HE PAS TRICHROME JONES/GMS

Cellularity Excellent Excellent Poor Poor

Mesangial M Poor Excellent Variable Excellent

Glom. Sclerosis Poor Excellent Excellent Good

Immune Cox. Poor Poor Variable Negative

Basement M. Poor Excellent Good Excellent

Fibrosis Poor Poor Excellent Excellent

Vascular hyaline Good Poor Good Negative

Thrombi Good Poor Good Variable

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H&E

Amyloid:• Silver negative• 8 micron sections

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Congo Red

Fibrillary GP

Diagnoses overlooked without IHC

1. Light chain-associated diseases2. AL amyloid3. Monoclonal immunoglobulin deposition disease4. Light chain cast nephropathy5. IgA nephropathy/Henoch–Shonlein purpura6. IgM nephropathy7. C1q nephropathy8. Antiglomerular basement membrane disease9. Humoral (C4d) transplant rejection10. Fibronectin glomerulopathy

IgG

IgA

Electron microscopic helpful, but not usually essential for the diagnosis1. Nephrotic syndrome2. Acute renal failure3. Chronic renal failure4. Renal disease in diabetes mellitus5. Renal disease in SLE6. Suspected rejection of a renal allograft7. Repeat specimen when the diagnosis has

been made

Electron microscopy is most likely to help

1. Microscopic Hematuria with normal renal function

2. Family history of renal disease.3. Asymptomatic proteinuria with normal renal

function.

Diagnoses overlooked without EM

1. Fibrillary /immunotactoid glomerulopathy2. Nail–patella syndrome3. Lipoprotein glomerulopathy4. Dense deposit disease5. Alport’s syndrome6. Thin basement membrane nephropathy7. Collagenofibrotic glomerulopathy

Thank You