1. CORE CURRICULUM IN NEPHROLOGY- AJKD
Agnes B. Fogo. MDs Francis
Kuwait
APPROACH TO RENAL BIOPSY

2. Sample Size (number of glomeruli)
Focal lesions involving a small number of glomeruli - minimum 25
Membranous glomerulonephritis - single
Transplant kidney biopsy diagnoses - minimum 7
For most light microscopic assessment - 8 - 10

3. Sample Location
(Juxtamedullary vs Cortical)
Subcapsular cortical samples have
overrepresentation of global sclerosis related to
aging/hypertension and non-specific scarring.
Juxtamedullary glomeruli are the earliest to be
involved with segmental sclerosis in focal
segmental glomerulosclerosis (FSGS). Preferred
location.

4. Microscopy preferrences
Native renal biopsies should include
Light microscopy (LM),
Immunofluorescence microscopy (IF),
Electron microscopy (EM).
For transplant biopsy
LM and IF are considered the standard
Repeat biopsies
only needing LM in many cases

5. Allocation of the biopsy tissue
EM - 1mm LM – largest IF - 3-4 mm

6. Fixatives
LM- Formalin (10% neutral buffered)
Paraformaldehyde
Bouin’s or Zenker’s (infrequent)
IF- Directly frozen,
Transport media such as Michel’s
(Tissue is stable at room temperature for mailing to
central laboratories in this media within one hour)
EM- Glutaraldehyde.

7. Handling of Tissue
No forceps, manipulate with thin wooden stick
to avoid crush artifact.
Avoid touching tissue with a LM or EM
fixative-contaminated scalpel or razor blade (this
contaminates the tissue for IF).

8. Inadequate tissue
IF tissue frozen for IF stains —
The remaining frozen tissue may be fixed in formalin
and processed for LM.
LM tissue fixed on the paraffin block-
1) EM study can be done by processing remaining tissue
on paraffin block
2) IF study can sometimes be done satisfactorily in
tissue that has not been in paraffin blocks too long

9. Staining and processing
1) LM- Dehydrated and placed in paraffin block,
and multiple serial sections are obtained and
stained.
Usual stains are-
• Hematoxylin & eosin,
• Periodic acid–Schiff (PAS),
• Silver methenamine (Jones)
• Masson trichrome.
Usual processing time – 5 hours

10. 2) IF-
Tissue for IF is surrounded with OCT compound
and frozen, and sections are produced
Stains - stained with
• Fluorescein-tagged antibodies against IgG, IgA, IgM,
• Complements C3 and C1q, and light chain.
• Complement product C4d may also be stained
(best on frozen tissue, with more technical difficulty
in staining on paraffin-block tissue)
Usual processing time – 2 hours

11. 3) EM-
EM tissue is processed and embedded in a
plastic, hard media and scout sections (so-called
thick sections) are obtained
Stain - with toluidine blue
Usual processing time – 2 days

12. Glomerulus Tubules
Interstitium Vascular disease
Assessment
Morphological Localisation

13. Category of Injury-
Active Versus Fibrosing
Active lesions
• Proliferation
• Necrosis
• Crescents
• Edema
• Active inflammation
( glomerulitis, tubulitis, vasculitis)
Fibrosing
•Glomerulosclerosis
•Fibrous crescents
•Tubular atrophy
• Interstitial fibrosis
•Vascular sclerosis

14. Stains Best assessment of
1. HE Cellularity and morphology
2 . PAS Basement Membrane
Mesangial matrix
3. Masson’s Trichrome Fibrosis

15. Stains Best assessment of
4. Jones Silver Basement Membrane
Mesangial matrix
5.Congo red Amyloid
6) MSB Fibrin

16. STAINING 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

18. • Green- Epithelial cells
a) Visceral- capillary walls i.e PODOCYTES
b) Parietal- Bowmans capsule & Proximal tubules
• Yellow – Endothelial cells lining capillary lumen
• Red – mesangial cells
• Blue – mesangial matrix

24. Lamina rara externa
Lamina rara interna
Lamina densa
Endothelial cell
Podocyte

25. Minimal change disease- LM

26. EM
Effacement of foot
processess
Condensation of
cytoskeleton to BM

27. Effacement of foot
processess

28. Membranous - LM
Thick capillary wall

29. EM

30. Effacement of foot
processess
Subepithelial
immune deposits
BM projection between
deposits

32. IF- mostly IgG global, rarely IgM and IgA

33. FSGS – LM
Sclerosis
Hyalinosis
Adhesion to Bowmans
capsule

34. FSGS

35. IF – IgM and C3

36. Thank You