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Education
How to do conjunctival and buccal
biopsies to investigate cicatrising
conjunctivitis: improving the
diagnosis of ocular mucous
membrane pemphigoid
CASE
A 79-year-old female presented with a 3-year history of trichia-
sis and red eyes (figure 1A). Multiple operations for entropion
and trichiasis had been carried out previously, and a conjunctival
biopsy investigating mucous membrane pemphigoid (MMP),
had been negative. Her right eye was blind due to retinal
detachment and glaucoma. Acuity was light perception in the
right eye (OD) and 6/12 in the left eye (OS). There was active
inflammation and bilateral inferior symblepharon, left upper lid
entropion and trichiasis. There were no systemic features of
MMP.
Conjunctival and buccal mucosa biopsies were undertaken.
Serum was tested for indirect inmunofluorescence. Conjunctival
and serum immunofluorescence were negative, but buccal
mucosa showed linear IgG and IgA staining along the basement
membrane zone (BMZ), consistent with MMP.
Figure 1 (A) The patient’s cicatrised inflamed left eye. (B) Taking a biopsy from the eye (not the same patient). If the speculum does not fit,
manually open the lids. (C) Taking a buccal biopsy. (D) Michel’s transport medium. (E) Direct immunofluorescence showing linear IgG, IgA along the
basement membrane zone.
530
Oral prednisolone 1 mg/kg/day on a tapering course over
6 weeks, and mycophenolate 1 g twice daily were commenced.
Nine months later, left upper lid entropion surgery was per-
formed with excellent results.
QUESTIONS (FOR ANSWERS SEE 537)
1. What investigations are needed to identify the cause of cica-
trising conjunctivitis?
2. Describe how to do conjunctival and buccal biopsies?
3. How would you manage the patient if you suspect ocular
MMP, but all immunopathology is negative?
Arturo E Grau,1 Jane Setterfield,2,3 Valerie P J Saw1,4
1
Cornea and External Disease Service, Moorfields Eye Hospital NHS Foundation
Trust, London, UK
2
Department of Oral Medicine, Guys Hospital, Guys and St Thomas’, NHS
Foundation Trust, London, UK
3
St John's Institute of Dermatology, Kings College London, London, UK
4
NIHR Biomedical Research Centre at Moorfields Eye Hospital and UCL Institute of
Ophthalmology, London, UK
Correspondence to Dr Valerie P J Saw, Cornea and External Disease Service,
Moorfields Eye Hospital, 162 City Road, London EC1V 2PD, UK; v.saw@ucl.ac.uk
Acknowledgements We gratefully acknowledge the support of John Dart in this
manuscript. We are also grateful for the expert advice of Professor Enno Schmidt
(Lubeck, Germany) regarding immunofluorescence testing. Valerie Saw is supported
by the NIHR Biomedical Research Centre at Moorfields Eye Hospital and the UCL
Institute of Ophthalmology.
Grau AE, et al. Br J Ophthalmol April 2013 Vol 97 No 4
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Education

Contributors VPJS initiated the concept of the paper, drafted and revised the Provenance and peer review Not commissioned; internally peer reviewed.
paper. She is guarantor. AEG drafted the paper. JS initiated the concept of taking To cite Grau AE, Setterfield J, Saw VPJ. Br J Ophthalmol 2013;97:530–531.
buccal biopsies in ocular patients who have no mouth symptoms or signs, provided
her own unpublished data for the paper discussion, and revised the draft paper. JD’s
patients were used for JS’s data, and he commented on the draft paper. Published Online First 19 January 2013
Competing interests None. Br J Ophthalmol 2013;97:530–531. doi:10.1136/bjophthalmol-2012-302963
Patient consent Obtained.

Grau AE, et al. Br J Ophthalmol April 2013 Vol 97 No 4 531

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Education

ANSWERS (FOR QUESTIONS SEE 530)
1. Ascertaining which of the causes of cicatrising conjunctivitis
in table 1 is responsible requires:
i. A thorough history of previous conjunctivitis (adeno-
viral, trachomatous, vernal) or skin disorders (Stevens–
Johnson, ezcema) and examination for typical features
of allergy, Sjögren’s, rosacea. Ask about extraocular
MMP symptoms.
ii. Referral to a specialist for biopsy of any skin or mouth
lesions
iii. Serum connective tissue disease screening
iv. Conjunctival biopsies in appropriate medium, for
instance, Michel’s medium (figure 1D) for direct
immunofluorescence (DIF). Consider a formalin biopsy
for any suspicion of neoplasia, sarcoidosis, or atopy.
v. A buccal mucosal biopsy is low risk and can be positive
in MMP when conjunctival biopsies are negative.
vi. If testing is available, take serum for IIF.
vii. Establish whether the conjunctival cicatrisation is pro-
gressive, or static.
2. If the patient’s eyes are severely inflamed, that is, active cicatris-
ing conjunctivitis, commence oral immunosuppression on the
day of the biopsy. Use an experienced immunofluorescence
laboratory, discuss with them beforehand and obtain Michel’s
medium.
Ideally, biopsy the upper bulbar conjunctiva where upper
eyelid coverage improves healing. Concerns about adverse con-
sequences in the event of glaucoma surgery have not been
observed. Avoid inferior bulbar conjunctiva because exposure,
blepharitis and dry eye can compromise healing. Avoid biopsies
in the conjunctival fornix, as these induce further scarring.1
After anaesthetic drops, a cotton bud soaked in anaesthetic
numbs the chosen biopsy site. If the conjunctiva is too con-
tracted, manually open the eyelids rather than using a specu-
lum. Spring scissors and non-crush forceps are used to take a
2–4 mm biopsy (figure 1C). Ensure that the size of the biopsy
is adequate, including the BMZ. Chloramphenicol ointment
is applied to the eye twice daily for 7 days, along with pred-
nisolone 0.5% or dexamethasone 0.1% drops, four times per
day for 4 weeks.
The main risk associated with a conjunctival biopsy is
inducing conjunctival inflammation and, potentially,
scarring.
This is avoided by commencing immunosuppression at
the time of biopsy, and a 4-week course of topical steroids.
When done carefully, conjunctival biopsy is well tolerated
and safe.2
Carrying out a buccal mucosal biopsy at the time of
taking conjunctival biopsies, regardless of whether patients
have oral disease or not, is simple and low risk. Inject 2 ml

Table 1 Differential diagnosis of cicatrising conjunctivitis

Static or very slowly progressive conjunctival scarring Progressive conjunctival scarring

1. Trauma: physical, chemical, thermal, radiation injury, artefacta 1. Neoplasia: squamous cell carcinoma, sebaceous cell carcinoma, lymphoma
2. Infection: 2. Mucous membrane pemphigoid (MMP).
Trachoma A. Ocular MMP/ocular cicatricial pemphigoid
Membranous streptococcal conjunctivitis B. Ocular MMP associated with other disorders:
Adenoviral conjunctivitis Linear IgA disease
Corynebacterium diphtheria Epidermolysis bullosa acquisita
Chronic mucocutaneous candidiasis Paraneoplastic MMP (anti-laminin 332 MMP)
Drug-induced ocular MMP
Stevens–Johnson syndrome
3. Allergic eye disease: 3. Other mucocutaneous and immunobullous disorders:
Atopic keratoconjunctivitis Mucocutaneous disorders: Lichen planus
Vernal keratoconjunctivitis Immunobullous disorders: Paraneoplastic pemphigus
4. Drug-induced conjunctival cicatrisation (DICC)*†
5. Mucocutaneous disorders:
Stevens–Johnson syndrome and TEN*
Graft-versus-host disease
Discoid and systemic lupus erythematosus‡
6. Immunobullous disorders:
Linear IgA disease*
Epidermolysis bullosa acquisita*
Bullous pemphigoid
Pemphigus vulgaris
Dermatitis herpetiformis
7. Systemic disease:
Rosacea
Sjögren’s syndrome
Sarcoidosis†
Scleroderma
Wegener’s granulomatosis
Inflammatory bowel disease
Ectodermal dysplasia*
Immune complex diseases
Porphyria cutanea tarda
Erythroderma ichthyosiform congenita
*A subset of patients with these diseases may develop autoantibody-positive progressive conjunctival scarring similar to ocular MMP.
†Associated with granulomatous conjunctival inflammation.
‡Rare cases can develop progressive scarring.

Grau AE, et al. Br J Ophthalmol April 2013 Vol 97 No 4 537

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Education
lignocaine with adrenaline using a 25 G needle, into the buccal
mucosa. Avoid the parotid duct, which opens opposite the 2nd
upper molar. Use a 4 mm dermal trephine (figure 1C) to
create a biopsy punch, then excise with scissors and forceps.
Place haemostatic pressure using gauze swabs for 1 min. No
suture is necessary, and antiseptic mouthwash is not usually
required.
3. If both the conjunctival and buccal biopsies are negative, and
the patient has no skin lesions for biopsy, and serology has
not shown any circulating BMZ autoantibodies, then provid-
ing other causes of conjunctival scarring (see table 1) have
been excluded, a clinical diagnosis of ‘autoantibody-negative’
MMP-like ocular disease can be made,3 based on evidence of
progressive scarring and typical clinical signs. These patients
are managed in a similar manner to ‘autoantibody-positive’
ocular MMP. Studies investigating these ‘autoantibody-
negative’ patients, regarding differences in clinical phenotype
and response to therapy, are currently being conducted.
DISCUSSION
A recent study investigating the incidence of cicatrising conjunc-
tivitis4 highlighted that 10% of ocular MMP cases did not have
a conjunctival DIF biopsy where there were no extraocular man-
ifestations supporting the diagnosis, and there was a median
delay of 7.5 months (up to 10 years) before the diagnosis was
made. Often clinicians are cautious about taking conjunctival
biopsies as they fear worsening the disease. We hope that this
description of how to take a conjunctival biopsy will help to
educate and encourage better assessment and investigation of
these patients.
Setterfield, Dart et al have conducted studies (unpublished) in
patients with pure ocular MMP (n=7), and have determined
that a biopsy from apparently healthy buccal mucosa for DIF
was positive in 3 / 5 (60%) patients, while conjunctival biopsies
538
were positive in 3 / 7 (43%) Additionally, in patients with pre-
dominantly ocular MMP (n=14) and a history of multisite
disease, a buccal biopsy was positive in 5 / 7 (71%) patients com-
pared with conjunctiva 6 / 10 (60%).
The presence of linear BMZ IgG and IgA in our case con-
firmed the diagnosis of MMP.5 Conjunctiva is fragile and difficult
to process for immunofluorescence; it is thought that this may
partly be the reason for only 50–86%6 7 of conjunctival DIF
biopsies being positive for MMP, in contrast to almost 100% of
biopsies from non-ocular sites including buccal mucosa, where
examination of the BMZ is easier given the nature of the tissue.8
In summary, the purpose of this article has been twofold: first,
to educate clinicians about how to do conjunctival biopsies and
avoid complications and second, to increase awareness about a
simple buccal mucosal biopsy, which can support the diagnosis in
the event of the conjunctival biopsies being negative.
REFERENCES
1 Wright P. Cicatrizing conjunctivitis. Trans Ophthalmol Soc UK 1986;105(Pt 1):1–17.
2 Frith PA, Venning VA, Wojnarowska F, et al. Conjunctival involvement in cicatricial
and bullous pemphigoid: a clinical and immunopathological study. Br J Ophthalmol
1989;73:52–6.
3 Saw VPJ, Dart JKG. Ocular cicatricial pemphigoid - Author Reply. Ophthalmology
2008;115:1640–1.
4 Radford CF, Rauz S, Williams GP, et al. Incidence, presenting features, and diagnosis
of cicatrising conjunctivitis in the United Kingdom. Eye (Lond) 2012;26:1199–208.
5 Chan LS, Ahmed AR, Anhalt GJ, et al. The first international consensus on mucous
membrane pemphigoid: definition, diagnostic criteria, pathogenic factors, medical
treatment, and prognostic indicators. Arch Dermatol 2002;138:370–9.
6 Bernauer W, Elder MJ, Leonard JN, et al. The value of biopsies in the evaluation of
chronic progressive conjunctival cicatrisation. Graefes Arch Clin Exp Ophthalmol
1994;232:533–7.
7 Thorne JE, Anhalt GJ, Jabs DA. Mucous membrane pemphigoid and
pseudopemphigoid. Ophthalmology 2004;111:45–52.
8 Setterfield J, Shirlaw PJ, Kerr-Muir M, et al. Mucous membrane pemphigoid: a dual
circulating antibody response with IgG and IgA signifies a more severe and persistent
disease. Br J Dermatol 1998;138:602–10.
Grau AE, et al. Br J Ophthalmol April 2013 Vol 97 No 4
 Downloaded from bjo.bmj.com on August 26, 2014 - Published by group.bmj.com

How to do conjunctival and buccal biopsies

to investigate cicatrising conjunctivitis:
improving the diagnosis of ocular mucous
membrane pemphigoid
Arturo E Grau, Jane Setterfield and Valerie P J Saw

Br J Ophthalmol 2013 97: 530-531 originally published online January

19, 2013
doi: 10.1136/bjophthalmol-2012-302963

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