Fig. 17.2 Retinal fragment in cell block (H&E x MP).
ocular pathologist. One wet-fixed slide is stained with a modi-
fied Papanicolaou stain, and the remaining fixed slides may be
saved unstained pending the need for special stains.
Cell Block Preparation
The sample is centrifuged for 5 minutes at 2500 rpm, and the
supernatant is decanted with a disposable pipette. The sediment
is overlaid with 1 mL of zinc formalin. Care is needed not to dis-
turb the sediment. After an hour the sediment can be carefully
loosened with an angle pick and gently teased onto a small tea
bag, which is then folded and placed into a cassette. The labeled
cassette is put into a container of 10% formalin. After histologic
processing, hematoxylin and eosin-stained cell block sections
are sent to the JSEI ocular pathologist for review. These sections
may also be used for special stains. The cell block preparation
technique is best when the sample is abundant.
A modification of this technique is used for very important
samples that are scant in quantity but for which sections would
be helpful for immunohistochemistry and special studies. In
this technique, the sample is processed manually in a 1.5-mL
microfuge tube. Centrifugation is performed for each change in
solution starting with formalin, through graded alcohols, up to
the clearing agent. Then the material is transferred to a metal
cassette where it is infiltrated with xylene and then paraffin on
a heating block. This manual technique is labor-intensive but
extremely effective at increasing the cellular yield for critical
Culture is the best method for bacterial identification, but
air-dried smears may be used as a preliminary demonstration of
bacteria. Papanicolaou-stained slides and the cell block are used
for identification of neoplastic or inflammatory processes. Cell
block sections demonstrate large tissue fragments (Fig. 17.2)
better than do cytospin preparations.
1. Perform standard conjunctival scraping.
2. Direct fluorescent antibody (DFA) technique uses a
monoclonal antibody and can be performed on a
routinely made smear.37 The preparation time is 15
minutes, after which the smear is ready for screening.
culture is the best method.38 The cultured
cells are grown on a coverslip for 48 hours and then
stained by DFA as in 2.
4. Giemsa stain is no longer used for
cation, because the DFA method is far more sensitive
To Detect Viral Cytopathic Effect (CPE)
1. Each viral group has CPE changes specific for that
family of viruses and, sometimes, specific for the
individual virus, e.g. herpes and cytomegalovirus
2. The specimen can be obtained by corneal scrape, con-
junctival scrape, retinal biopsy, or subretinal fluid.
3. After culturing the specimen, viral-specific CPE is
noted and can be confirmed by DFA. If CPE is subtle
or not found, DFA can indicate early subvisual
Anatomy and Histology
It is useful to consider four compartments from which samples
may be attained, with the first three compartments referred to as
the adnexae: eyelid, conjunctiva, orbit, and eye.
The eyelids are bifaced, functioning as protective skin exter-
nally and as lubricating conjunctival membranes on their inner
surfaces. As such, they are affected by lesions identical to those
of the skin and mucous membranes elsewhere. Unique to the
area are the lacrimal and the meibomian glands. The lacrimal
glands are histologically tubuloalveolar serous glands, and
function to produce tears. The meibomian glands are special-
ized sebaceous glands.
The conjunctiva is a thin membrane, which conjoins the eye
and eyelids. The epithelial lining varies according to location.
The conjunctiva nearest the cornea has a specialized squamous
epithelium but relatively few goblet cells while epithelium in
the recesses of the eyelids is covered by more cuboidal shaped
epithelium with numerous goblet cells. The conjunctiva secretes
an array of mucins and there are numerous accessory lacrimal
glands in the eyelid that lubricate the eye.
The bony orbit has four walls forming a pear shape, open on
opposite sides. The walls are called the roof, floor, and lateral and
medial walls and are related anatomically to the frontal, maxil-
lary, ethmoid, and sphenoid sinuses. Therefore, diseases affect-
ing the sinuses can involve the orbit and vice versa. Contained
within the orbital cavity are the lacrimal glands, extraocular
muscles, arteries and veins to the area, various nerves, fibrofatty
tissue, optic nerve, and the eye. For a more graphic understand-
ing, readers are referred to any standard textbook of anatomy.
The wall of the eye (Fig. 17.3) is divided into an outer coat
(sclera and cornea), a middle coat or uvea (choroid, ciliary body,
and iris), and the inner coat (retina). Of the outer coat, only the
cornea participates in vision and therefore must be transparent
and avascular. The ciliary body is intimately associated with the
lens and is responsible for its support and contour. At the back
of the eye, the retina is thickest (ten layers) and is the functional
peripheral neurologic portion of sight.
The optic nerve is an extension of the brain and ends in the
retina. The meninges covering the brain are contiguous with the
optic nerve as it enters the bony orbit.