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    Eye biopsies and local excisions


    A range of lesions and diseases affect the eye that may require biopsy or local excision.1-5 A separate protocol is provided for enucleation and exenteration specimens.

    Eyelids may be affected by lesions seen in other skin specimens such as cysts, papillomas, pigmented, inflammatory, vascular and neoplastic lesions.2 A chalazion (or meibomian cyst) is a common, often recurrent, cystic lesion caused by blocked meibomian glands in the eyelid. Malignant tumours of the eye such as retinoblastomas, malignant melanomas and carcinomas of the conjunctiva, eyelid and lacrimal gland as well as sarcomas and lymphomas also may be found in eye specimens.

    Corneal small biopsies, epithelial scrapings, corneal buttons, corneal endothelial keratoplasty and donor corneal specimens may all be received in the laboratory.2

    Record the patient identifying information and any clinical information supplied together with the specimen description as designated on the container. See overview page for more detail on identification principles.

    • No
      • Non-routine fixation (not formalin), describe.
    • Yes
      • Special studies required, describe.
      • Ensure samples are taken prior to fixation.

    Note the importance of aseptic technique in handling fresh tissues (see overview for more information).

    • Consider if microbiology is needed and check if a separate sample has been submitted or if the one received needs to be divided before proceeding
    • Frozen sections (e.g. for sebaceous carcinoma)
    • Cytology(imprints may be useful if there is a question of ocular surface squamous neoplasia or Vitamin A deficiency)
    • Immunofluorescence (all ciciatrising conditions)
    • Flow cytometry (for lymphoma)
    • Polymerase chain reaction (PCR) may be indicated for viral infection investigation.5
    • Electron microscopy may be required on corneal specimens when dystrophy is suspected. Transfer an axial strip to glutaraldehyde if required.2 It is always preferable to have the surgeon divide the specimen. Ensure EM fixative is available in theatre.

    See general information for more detail on specimen handling procedures.

    Inspect the specimen and dictate a macroscopic description.

    External Inspection

    Orientate and identify the anatomical features of the specimen.

    Record additional orientation or designation provided by operating clinician:

    • Absent
    • Present
      • Method of designation (e.g. suture, incision)
      • Featured denoted

    Photograph the intact specimen if required.

    Describe the following features of the specimen:


    Record as stated by the clinican

    • Conjunctival biopsy –refer to head and neck small biopsy protocol
    • Corneal
      • Button/disc
      • Biopsy
      • Donor corneal-scleral tissue
    • Eyelid
    • Partial thickness
      • Small biopsy
      • Incisional biopsy
      • Skin ellipse
    • Full thickness/pentagonal wedge2
    • Iridectomy
    • Local resection of ciliary body and choroid
    • Endo-resection -*treat as a cytology specimen.

    Record the volume of fluid (ml), describe any tissue fragments present and spin down for preparation onto a slide (e.g. cytospin) or for processing as a cell block.2

    Anatomical components (more than one may apply) and specimen dimensions (mm)1-5

    Describe and measure the components present, as follows:

    • Measure the maximum dimension (i.e. diameter if circular)
    • Note any variation in thickness
    • Note any defects, opacity, ulceration or scarring
      • Describe distribution, shape, colour and measure each in maximum dimension
    • Note any neovascularisation
    • If the specimen is a ring of remnant (donor) corneal-scleral tissue, also measure the diameter of the circular defect (inner diameter). Note the presence of a trabecular meshwork, ciliary muscle or other relevant features of the specimen.
    • Measure in three dimensions
    • Note the presence of the lid margin or tarsal conjunctiva2
    • Note the presence of any eyelashes
    • Measure in three dimensions
    • Measure the vertical thickness of the skin and conjunctival surfaces

    A dissecting microscope may be required to examine corneal specimens clearly.

    Back illumination and viewing against both dark and light backgrounds will assist in identifying abnormalities such as opaque, ulcerated or scarred areas and whether the specimen is full or partial thickness. Note that partial-thickness corneal tissue will have an irregular outline and is more likely to fold and roll up.5

    These specimens must be processed with full knowledge of the clinical presentation. Resection specimens require orientation by the surgeon.

    • Record the anatomical structures present
    • Measure in three dimensions (pupil margin to peripheral/scleral margin, circumference to circumference margin and thickness).
    • Note the presence of tumours, iris atrophy (using back illumination), previous biopsy sites, neovascularisation, ectropion uveae or entropion uveae.

    These specimens must be processed with full knowledge of the clinical presentation. Resection specimens require orientation by the surgeon.

    • Record the anatomical structures present e.g. iris, ciliary body and sclera
    • Measure in three dimensions
    • Paint the surgical margins with ink

    Specimen integrity

    • Intact
    • Disrupted


    Various techniques are used for ophthalmic specimens:1-5

    Eyelid –partial thickness (small biopsy, skin ellipse, incisional biopsy)

    • Specimens with orientation markers should be painted with ink along the surgical margin(s)2
    • Refer to skin protocol for the dissection of skin ellipses and incisional biopsies2
    • Specimens with orientation markers should be painted with ink to identify the surgical margin(s).2 This is best achieved by painting the lateral and medial halves of the specimen with contrasting colours.
    • Serially section the specimen at 2-3 mm intervals5
    • Refer to skin protocol for the dissection of skin ellipses and incisional biopsies2
    • Handle corneal specimens with care to avoid damage to the anterior and posterior surfaces2,5
    • Small corneal biopsies do not required dissection2
    • With the epithelial (convex) surface down (i.e. cutting from the endothelial surface), bisect corneal buttons/discs through the area of interest along the longest axis (at right angles to the direction of wounds and scars).2,5
    • However, if area of interest is very small, cut just to one side of the abnormality so it is entirely within one piece of tissue (to prevent loss during trimming of the block).
    • Ensure that instructions are communicated for tissue to be embedded on edge so the full thickness of each piece is demonstrated microscopically.
    • Submit all tissue for processing.
    • Special stains will be required; routinely Periodic acid Schiff’s and others depending on the clinical history.

    Where minimal tissue is available, a 1-2mm central strip from the specimen may be sufficient for routine histology. This will allow peripheral segments to be available for other tests such as electron microscopy or polymerase chain reaction (PCR) for viral infection investigation.

    • If cornea is present, dissect into three parallel sections2
    • Where there is a central defect present and only a scleral rim remains, section into 4mm segments radially around the circumference2

    Paint the circumferential resection margins and slice at sequential intervals around the circumference so each section contains a pupil and peripheral margin.

    • Paint the scleral/iris resection margins and submit in separate cassettes
    • Serially slice in the anteroposterior plane

    Internal Inspection

    Describe the cut surface appearance including the following items:


    • Absent
    • Present
    • Number; if more than one tumour, designate and describe each tumour separately

    Tumour size (mm)

    • Maximum dimension

    Tumour growth pattern

    • Endophytic
    • Exophytic

    Non-tumour lesion, describe

    • Colour
    • Texture/consistency
    • Borders
    • Relationship with normal tissue
    • Haemorrhage
    • Necrosis

    Separately submitted

    • For each specimen container, record specimen number and designation
    • Collective size of tissue in three dimensions (mm)
    • Number of grossly identified lymph nodes submitted
    • Maximum diameter of each (mm)

    Photograph the dissected specimen if required.

    Note photographs taken, diagrams recorded and markings used for identification.


    Dissect the specimen further and submit sections for processing according to the illustrations provided.

    Submit representative sections of:

    • Small biopsies are processed whole. Transfer directly to cassettes for processing.
    • Submit both halves of corneal buttons/discs for processing.
    • Submit all sections of donor corneal-scleral specimens.
    • Circumferential margins may be processed in a separate cassette.
    • Ensure that instructions are communicated for tissue to be embedded on edge so the full thickness of each piece is demonstrated microscopically.
    • Tumour with nearest resection margin
    • Circumferential margins in separate cassette
    • Tumour with nearest resection margins
    • Shave margins

    If received, submit all lymph nodes and identify the site of each.

    Lymph nodes may be received, where melanoma or retinoblastoma is suspected, to investigate extrascleral metastases as there are no intraocular lymphatics.1

    Record details of each cassette.

    An illustrated block key similar to the one provided may be useful.

    Block allocation keys

    Cassette id Site No. of pieces
    A Eye biopsy, all tissue  
    Cassette id Site No. of pieces
    A Tumour and resection margin(s)  
    B Circumferential margins  


    Assoc Prof Sonja Klebe for her contribution in reviewing and editing this protocol.


    1. Lester SC (ed). Manual of Surgical Pathology, Saunders Elsevier, Philadelphia, 2010.
    2. Mudhar HS, Irion L. Tissue pathways for non-neoplastic ophthalmic pathology specimens. The Royal College of Pathologists; London, 2015.
    3. Menocal NG, Ventura DB and Yanoff M. Routine processing of ophthalmic tissue for light microscopy. Am J Med Techno. 1977; 43(2):156-160.
    4. Torczynski E. Preparation of ocular specimens for histopathologic examination. Ophthalmology. 1981; 88(12):1367-1371.
    5. Ford AL, Mudhar HS, Farr R, Parsons MA. The ophthalmic pathology cut-up–Part 2. Current Diagnostic Pathology. 2005;11(5):340-8.

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