Tissue staining (chromoscopy)of the gastrointestinal tract

M Brian Fennerty MD

Tissue staining, or chromoscopy, is an adjunctive endo-scopic technique using chemical agents applied to the

gastrointestinal mucosal surface to identify specific epitheliaor to enhance the mucosal surface characteristics of the gas-trointestinal epithelium. Chromoscopy is performed to aidin the recognition of subtle lesions (ie, polyps), or to allow -directed targeting of biopsies (ie, sprue or Barrett’s esopha-gus) to increase the yield of endoscopic diagnostic accuracy.

There are four endoscopic tissue staining techniques: vi-tal staining, contrast staining (chromoscopy), reactive stain-ing and tattooing (1-4). Vital staining is the use of an agentthat is absorbed by the gastrointestinal epithelium, allowingthe identification of a characteristic epithelium. An exam-ple of vital staining is the use of methylene blue to identifygastric intestinal metaplasia. Contrast staining, or chromos-copy, is the use of an agent to accentuate the surface topogra-phy of the gastrointestinal mucosa, allowing the identifica-

tion of subtle changes that otherwise may be unrecognized.An example of chromoscopy is the use of indigo carmine toidentify flat colonic lesions. The term ‘chromoscopy’ is usedinterchangeably with ‘tissue staining’ to refer to all forms ofthese tissue staining techniques. Reactive staining is the useof agents that identify chemical reactions within the gastro-intestinal tract. For instance, Congo red turns black in thepresence of acid and has been used to identify acid-secretingportions of the stomach. Tattooing is a technique that useseither short-acting or permanent agents such as India ink tomark a specific mucosal site for longitudinal study.

Chromoscopy is not a new technique. Tissue staining wasperformed by pathologists for many years before the develop-ment of the endoscope (4). These techniques were adaptedto endoscopic practice shortly after the introduction of rigidand fibreoptic instruments. Tissue stains are usually ‘sprayed’onto the mucosal surface during endoscopy, but have also

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Division of Gastroenterology, Oregon Health Sciences University, Portland, Oregon, USACorrespondence: Dr Brian Fennerty, Oregon Health Sciences University/PV 310, 3181 SW Sam Jackson Park RoadPortland, Oregon 97201-3098, USA. Telephone 503-494-8577, fax 503-494-7556, e-mail Fennerty@OHSU.edu

11th INTERNATIONAL COURSE ON THERAPEUTIC ENDOSCOPY

MB Fennerty. Tissue staining (chromoscopy) of the gastroin-testinal tract. Can J Gastroenterol 1999;13(5):423-429. Tissuestaining, or chomoscopy, is used as an adjunctive technique dur-ing gastrointestinal endoscopy. Chemical agents are applied to thegastrointestinal mucosal surface to identify specific epithelia or toenhance the mucosal surface characteristics of the gastrointestinalepithelium. This aids in the recognition of subtle lesions (ie, pol-yps) or allows directed targeting of biopsies (ie, sprue or Barrett’sesophagus) to increase the yield of endoscopic diagnostic accu-racy. The four endoscopic tissue-staining techniques in use are vi-tal staining, contrast staining (chromoscopy), reactive stainingand tattooing. Some of the agents used for endoscopic tissue stain-ing and the uses of chromoscopy in identifying pathology of theesophagus, stomach, small bowel and colon during endoscopy arediscussed.

Key Words: Chromoscopy; Endoscopy; Tissue staining

Coloration tissulaire (chromoscopie) des voiesdigestivesRÉSUMÉ : La coloration tissulaire, ou chromoscopie, est utilisée commetechnique d’appoint durant l’endoscopie gastro-intestinale. Des agentschimiques sont appliqués à la muqueuse gastro-intestinale pour identifierles épithélia spécifiques ou pour rehausser les caractéristiques de surface dela muqueuse de l’épithélium gastro-intestinal. Cette technique facilite lareconnaissance des lésions discrètes (par exemple, polypes) ou permet unciblage plus direct des biopsies (par exemple, sprue ou œsophage deBarrett) afin d’augmenter la précision diagnostique de l’endoscopie. Lesquatre techniques de coloration tissulaire endoscopique en usage sont lacoloration vitale, la coloration de contraste (chromoscopie), la colorationréactive et le tatouage. On présente ici certains des agents utilisés pour lacoloration tissulaire endoscopique et les emplois de la chromoscopie pourl’identification des pathologies de l’œsophage, de l’estomac, du grêle et ducôlon durant l’endoscopie.

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been given orally and rectally before the procedure. Despitechromoscopy’s many uses and advantages, this endoscopictechnique is not often used by North American endo-scopists, and few endoscopists have been formally trained inchromoscopy (2).

Some of the agents used for endoscopic tissue stainingand the uses of chromoscopy in identifying pathology of theesophagus, stomach, small bowel and colon during endo-scopy are discussed.

TISSUE STAININGMany different chemical compounds have been used as tis-sue staining agents (Table 1). Vital or absorptive stains in-clude Lugol’s solution, indocyanine green (IG), methyleneblue and toluidine blue. Lugol’s solution is an inexpensive,widely available, nontoxic mixture of iodine and potassiumiodide forming a compound iodine solution. First describedby the Parisian Jean Guillaume Auguste Lugol in the 1800s,Lugol’s solution stains the normal nonkeratinized squamousepithelium of the normal esophagus. Lugol’s solution istaken up by squamous epithelium because of glycogen’s af-finity for iodinated agents. Once absorbed by the squamous

epithelium, Lugol’s solution results in a characteristicgreen-brown colour, the intensity of which is in part depend-ent upon the amount of glycogen present. The absence ofstaining with Lugol’s solution can be used to identify abnor-mal epithelium such as inflammatory squamous epithelium,neoplastic squamous epithelium or metaplastic epitheliumwithin the esophagus. Lugol’s solution appears to be safe, al-though in one study a patient later found to be allergic to io-dine developed bronchial spasm following its use in theesophagus (5). Lugol’s solution should, therefore, be usedcautiously in patients with reported iodine sensitivity. In thesame study, three of 46 individuals also reported transientheartburn following installation of a 50% solution into theesophagus (5).

IG is composed of 1 to 2 µm dimers and polymers of a car-bon, nitrogen and sulphur compound in solution. IG(5 mg/kg) is usually given as an infusion of 5% albumin 15 to30 mins before visualization of the liver. Following infusion,IG is taken up by normally functioning hepatocytes, allow-ing one to differentiate normal from abnormal hepatocellu-lar function. Thus, IG enhances the identification ofdiseased hepatic tissue for directed biopsy during open orlaparoscopic visualization of the liver. IG has also been usedas a colonic tattooing agent, but is less widely studied thanthe more commonly used and widely available endoscopictattoo, India ink.

Methylene blue (methylthionine chloride) is a nontoxiccarbon-based chemical agent that reversibly stains activelyabsorbing epithelium such as the epithelium of the small in-testine and colon. Methylene blue is not absorbed by thesquamous epithelium of the esophagus or the columnar epi-thelium of the stomach. However, metaplastic epitheliumwith absorptive characteristics such as intestinal metaplasiain the stomach (gastric intestinal metaplasia) or esophagus(Barrett’s esophagus) is capable of absorbing the stain.Methylene blue can be used to identify this metaplastic ab-sorptive epithelium or gastric metaplasia of the duodenalbulb in the absence of uptake when used in the small intes-tine (6,7).

Following application of 10 to 50 cm3 of a 5% to 10% so-lution of methylene blue either orally preprocedure or duringendoscopy by spraying, uptake of the stain occurs within 2 to3 mins, which is then resistant to vigorous washing or lavage(Figure 1). The stain fades over the next 15 to 30 mins butpersists, in most patients, for up to 12 to 24 h until the stain islost, secondary to renal excretion or cellular loss. The stainis preserved in frozen section biopsy specimens but lost dur-ing permanent tissue fixation with formalin and alcohol.Methylene blue results in blue discoloration of the urine andstools for the next 24 to 48 h.

In order for methylene blue to come into contact with ab-sorptive epithelium and allow staining, surface mucus mustfirst be removed. Proteinase (pronase) and mucolytic agents(N-acetylcysteine [Mucomyst, Bristol Laboratories, Evans-ville, Indiana]) have been used to remove the mucous layer.N-acetylcysteine contains sulphydryl groups that result inthe disruption of disulphide bridges of the mucous layer. This

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TABLE 1Tissue stains

Class Use

Tattooing agents

India ink For permanent marking of a mucosal site forrelocalization at the time of surgery orendoscopy. Used in the esophagus,stomach and colon. Safe and without sideeffects.

Indocyanine green Shorter-duration tattooing agent than Indiaink.

Absorptive stains

Lugol’s iodine Stains normal glycogen-containing squamousmucosa in the esophagus, allowingrecognition of abnormal squamousepithelium (dysplasia) or metaplasticepithelium (Barrett’s esophagus).

Methylene blue andtoluidine blue

Stains the absorptive epithelium (small boweland colon), allowing the identification ofmetaplastic epithelium in the esophagus(Barrett’s esophagus) and stomach (gastricintestinal metaplasia). Also can identifygastric metaplasia (negative stain) in theduodenal bulb.

Contrast stains

Indigo carmine andcresyl violet

Accentuates mucosal topography, allowingrecognition of abnormal small bowel(sprue) and colonic mucosa (inflammatorybowel disease, polyps).

Reactive stains

Congo red Identifies acid-secreting portions of thestomach postoperatively; documentsachlorhydria.

Phenol red Identifies alkaline areas of the stomach.

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action destroys the glycoproteins critical to the integrity ofthe mucous cap, resulting in disintegration of the mucouslayer. N-acetylcysteine can be given orally or sprayed di-rectly onto the mucosal surface before application of methyl-ene blue. However, N-acetylcysteine has a foul taste and asulphurous odor, making oral ingestion unpleasant, and thisform of application is rarely used.

Toluidine blue stains neoplastic cell nuclei. This basicmetachromatic dye has been postulated to either have a tro-pism for the rapidly proliferating cells, characteristic of neo-plastic tissue, or stain related to its absorption based ondecreased cellular adhesion and ability to penetrate what

was previously a ‘tight junction’. Toluidine blue’s most com-mon application, as a tissue stain, has been in the identifica-tion of neoplastic oral squamous epithelium when used as a1% solution, following a 1% application of acetic acid to re-move surface debris. No toxicity or side effects have been ob-served with this agent when used for this purpose.

CONTRAST STAINS (CHROMOSCOPY)Contrast stains (chromoscopy) include cresyl violet and in-digo carmine. Cresyl violet pools in the margins of the colo-nic pits, resulting in a ‘stain’, highlighting the topography ofthe colonic epithelium. While less commonly used during

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Tissue staining of the GI tract

Figure 1) Prominent focal (Top left, top right, bottom left) and subtle diffuse (Bottom right) staining, following application of 10% N-acetylcysteineand a 5% solution of methylene blue

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chromoscopy than indigo carmine, it appears to be an ef-fective, nontoxic agent when used as a colonic contrast stain.

Indigo carmine is a blue nontoxic dye (indigo) derivedfrom plants combined with carmine, a compound of cochi-neal and alum, which forms a red colouring agent. Indigocarmine is not absorbed and percolates across the surface ofthe epithelium following application. The stain collects inthe sulci and grooves of the mucosa, highlighting the topog-raphy of the stained epithelium. Indigo carmine has beengiven as 100 mg of a powder in a capsule, or sprayed onto themucosa as a 0.1% to 2% solution. No side effects or toxicityof indigo carmine has been described.

REACTIVE STAININGReactive staining is the use of pH-dependent dyes that canbe used to highlight acid-secreting or alkaline-associatedepithelia. Congo red, in the presence of acid (pH less than3), decomposes, resulting in a colour change from red to ablue-black pigment. Congo red is almost always used with asecretagogue such as pentagastrin to detect acid-secretingepithelium. Pentagastrin is used to insure adequate acid se-cretion to enhance the accuracy of the test. Pentagastrin(250 mg) is usually given intramuscularly 30 mins before theprocedure, and then surface acid is neutralized with a 0.5%sodium hydroxide solution before spraying 10 to 50 cm3 ofCongo red in a 0.3% to 0.5% solution directly onto the sur-face epithelium in the stomach or duodenum. Congo red isnontoxic and without side effects.

Phenol red is a yellow agent that turns red in the presenceof alkali. This agent has been used to depict areas with highHelicobacter pylori concentration, as well as atrophic gastritis.Phenol red, as is the case with Congo red, appears to be non-toxic and without side effects.

TATTOOING AGENTSIndia ink has been the most widely used tattooing agent. In-dia ink is a colloidal suspension of carbon particles. The car-bon is derived from incomplete combustion of petroleumproducts in an oxygen-depleted system. When this carbonpowder is combined with dilutants, stabilizers and surfac-tants such as alcohol, shellac, phenol and ammonia, Indiaink is formed. While there are numerous manufacturers ofthis product, all of the products contain the amorphous car-bon particles characteristic of India ink.

Carbon tattooing has been used for decades to tattoo theskin without toxicity, and India ink tattoos of the gastroin-testinal tract appear similarly to be safe. Tattooing of the co-lon with a black tattoo dye was first attempted by Knoerchildin 1962 (8), using a rheostat inserted through a rigid proctos-cope. Tattooing via an injection catheter at the time of flexi-ble endoscopy was described in 1975 (9). Since then, morethan 200 cases of tattooing of the esophagus, stomach andcolon have been described in the literature (10-12). The tat-too obtained with this technique appears to be long lasting,if not permanent. Although ‘histological’ complications offat necrosis, inflammation, abscess and focal peritonitis havebeen described, they have not resulted in any clinical se-quela, and these pathological ‘complications’ were only ser-endipitous findings. Furthermore, long term safety appears tobe a nonissue because there is little or no inflammatory re-sponse to the amorphous carbon particles remaining in thelamina propria and submucosa (10-12).

India ink, diluted in a one to 10 ratio with sterile saline,can be prepared in a 5 mL high pressure liquid chromatogra-phy vial and then autoclaved. Alternatively, the solutioncan be drawn through a millipore filter needle. The lattertechnique removes a substantial portion of the carbon,which may affect the quality and persistence of the stain. Ta-ttoos with India ink are usually applied by injecting less than1 mL of the sterilized solution with a sclerotherapy needle.After the sclerotherapy needle is purged with 0.75 to1.25 mL of India ink, a 0.3 to 0.5 mL aliquot of solution is in-jected tangentially to provide a small bleb in the mucosa(Figures 2,3). Care should be taken to avoid deep injection,which stains the peritoneum and may obscure the surgeon’sview, or luminal spillage, which makes further endoscopicinspection somewhat difficult.

Methylene blue, indigo carmine and toluidine blue havealso been used as tattooing agents with little success, relatedto their transient staining qualities. However, IG appears tobe longer lasting, although it does not result in a persistentstain, limiting its applicability as a tattooing agent.

TISSUE STAINING OF THE ESOPHAGUSSquamous epithelium: In the esophagus, Lugol’s solution ofpotassium iodine can be used to differentiate normal squa-mous epithelium from abnormal inflammatory or neoplasticsquamous epithelium, or metaplastic epithelium such as thecolumnar epithelium of Barrett’s esophagus (Figure 4). Thelargest use of Lugol’s solution is in identifying dysplastic ormalignant squamous epithelium. While large malignancies

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Figure 2) Esophageal India ink tattoo used before studying the thermalablation of Barrett’s esophagus

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of the esophagus are readily identifiable, early lesions or thefull extent of the malignant involvement is not always evi-dent at the time of endoscopy. Lugol’s solution can be usedto identify early subtle neoplastic esophageal processes, aswell as stage more accurately the extent of mucosal involve-ment. When identifying esophageal cancer, 20 to 50 cm3 of a1% to 50% solution was used.

While Lugol’s iodine has not been studied as extensivelyfor this purpose, it should also be able to detect, by negativestaining, inflammatory esophageal epithelium that has fewerintact glycogen-containing cells. Similarly, small, previouslyunrecognized areas of columnar epithelium can be observedas a negative stain following installation of Lugol’s solutioninto the esophagus. Woolf and co-workers (13) determinedthat the sensitivity of Lugol’s solution was 89% and thespecificity 93% when used for this purpose. Lugol’s solutionhas also been used alone or in conjunction with Indigo car-mine and magnification endoscopy to increase the recogni-tion of short segments of Barrett’s esophagus. This haspotential screening and therapeutic implications, and maybecome more applicable now that endoscopic techniques arebeing actively investigated to attempt to eradicate Barrett’sesophagus.

More recently, the absorptive characteristics of methyl-ene blue have been used to identify the specialized columnarepithelium of Barrett’s esophagus, in order to increase the ac-curacy of endoscopic biopsies (14). Canto and colleagues(14) determined that a 0.5% solution of methylene blue (fol-lowing installation of 10% solution of N-acetylcysteine) hadan accuracy of 95% in detecting specialized columnar epi-thelium of Barrett’s esophagus. The controls in that studyhad a diagnostic yield of 45%, implying that Barrett’sesophagus was routinely missed. Ninety-five per cent of

stained specimens demonstrated specialized columnar epi-thelium versus 3.5% of nonstained biopsies. This stainingwas reproducible at eight weeks, and dysplasia was stained aswell. The risk of dysplasia in a stained biopsy specimen hadan odds ratio of 17.7 (P=0.0004). This technique, which iseasily performed, deserves further scrutiny because it may al-low improved accuracy of endoscopic screening and surveil-lance of Barrett’s esophagus.

Tattooing has also been used in the esophagus. India inktattoos have been applied to insure precision of measure-ment, relocalization and rebiopsy of specific sites. This is, inpart, in response to the imprecision of standard endoscopicmeasurements. India ink tattooing of the esophagus was firstreported in 10 patients as part of a Barrett’s esophagus eradi-cation trial using bipolar probe therapy (BICAP, CirconACMI, Connecticut) (15). Since then, Shaffer and col-leagues (12) studied 19 patients after 0.1 mL of India ink, di-luted in a one to 10 ratio with normal saline, was used todemarcate the proximal extent of Barrett’s esophagus.Follow-up at three, nine, 15, 24 and 36 months revealed a95% persistence of the tattoo at three months. All 15 pa-tients evaluated at 36 months had persistence of good to ex-cellent tattooing with this agent. There were nocomplications or side effects, and in nine cases biopsiesshowed persistence of the carbon particles without inflam-matory response.Stomach: Methylene blue has been studied extensively as ameans of staining gastric intestinal metaplasia (3,4,6,16).Once surface mucus is removed with a mucolytic agent andmethylene blue is applied, islands of methylene blue stainingepithelia are easily identified. A variety of staining formshave been described: focal versus diffuse and subtle versusprominent. The more prominent stains correlate to a greater

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Tissue staining of the GI tract

Figure 3) Proximal and distal India ink tattoos in the colon Figure 4) Unstained mucosa (arrow) of the esophagus followingapplication of Lugol’s solution. Biopsy revealed Barrett’s mucosa thatwas otherwise unrecognized

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‘density’ of goblet cells seen on pathological examination.Theoretically, staining the gastric intestinal metaplasia withmethylene blue should allow quantification of the amount ofgastric involvement by intestinalized epithelium, as well aslongitudinal follow-up to establish the natural history of thelesion as well as, for example, its response to anti-helico-bacter therapy. Most recently, methylene blue has been usedin the stomach to map intestinal metaplasia of the cardia(16). In that study, the use of methylene blue increased therecognition of this lesion.

Congo red and phenol red can be used to map acid-secreting and alkaline areas of the stomach. In the past,Congo red was used to assess the adequacy of a surgical va-gotomy. Congo red has also been used in combination withmethylene blue to identify dysplasia in the stomach(‘bleached areas’ not stained with methylene blue or Congored). Phenol red has been used to identify alkaline areaswithin the stomach. Dense colonization of H pylori can beidentified by this technique because the local production ofammonia by the organisms results in a local alkaline pH.Small bowel: The absorptive dye methylene blue has beenused to identify metaplastic gastric epithelium in the duode-nal bulb by negative staining (7). The most common clinicaland research application of tissue staining in the small bowelhas been the use of contrast agents in the identification ofceliac sprue. It is estimated that the prevalence of sprue is ashigh as one in 230 people of European decent. The spectrumof illness ranges from the asymptomatic individual to a life-threatening malabsorptive process. The diagnosis of sprueremains histological, but certain endoscopic markers such asscalloping of the mucosa, loss of duodenal folds and a mosaicmucosal appearance increase the likelihood of sprue beingpresent. The sensitivity and specificity of these endoscopicfindings are 94% and 92%, respectively. However, these en-doscopic appearances may be subtle, absent or unrecognized.Dye spraying has been used to facilitate the delineation ofthis surface morphology (17,18). Indigo carmine as a con-trast agent was used as early as 1976, and in a more recentstudy was significantly more effective when combined with amagnifying endoscope than standard endoscopy in identify-ing partial atrophy characteristic of sprue (17). This tech-nique was 91% accurate versus 9% with a standardendoscope without dye spraying (P<0.01). Methylene bluehas also been used as a small bowel contrast agent. In avideo-endoscopic study, the kappa value for identifying theloss of folds with this technique was 0.59, and for scallopingwas 0.76.Colon: Tissue staining has been best studied in the colon as ameans of identifying small or flat neoplastic polyps that oth-erwise would go unrecognized (19-22). Tissue staining hasalso been used in the surveillance of ulcerative colitis(23-24), and in tattooing of polypectomy sites for surgical re-localization, once a polyp is found to be malignant (10). Italso has other potential applications.

Identifying whether a patient is an adenoma former hasclinical implications for further endoscopic surveillance rec-ommendations, yet endoscopists are aware that small polyps

may be difficult to identify and are missed frequently at thetime of colonoscopy. Furthermore, some investigators havenoted that small, flat adenomas (which may be very difficultto detect) have an increased incidence of dysplasia and ofmalignancy. Thus, enhanced recognition of small lesionsmay be clinically relevant.

Numerous studies have established that chromoendo-scopy enhances the visualization of diminutive colonic le-sions. Mitooka and colleagues (20) compared indigocarmine and magnifying endoscopy with standard endo-scopy, and demonstrated enhanced detection as well as visu-alization of flat lesions (7.7% versus 1.8%). Thus,chromoendoscopy was shown for the first time not only toenhance the appearance of recognized lesions, but also to in-crease the yield of detection. Matsumoto et al (23) also de-tected four cases of minute (2 to 4.5 mm) flat lesions thatcontained invasive cancer, detected by chromoendoscopy,with a 1% to 2% solution of indigo carmine. Because of thisobservation, they have routinely used dye spraying of the en-tire colorectal mucosa since 1990. Others have also demon-strated that the accuracy of detection of small lesions isincreased with dye spraying (16). More recently, Axelradand colleagues (22) used dye spray combined with magnify-ing endoscopy to differentiate adenomatous polyps from hy-perplastic polyps (22). The overall accuracy of thistechnique was 82%, based on differences in surface charac-teristics of the lesions observed with dye spraying. Cresyl vio-let has also been used as a chromoendoscopy agent in similarstudies of the colon.

Dye spraying or chromoendoscopy has also been used inulcerative colitis to quantify inflammation and detect earlyneoplasia (23,24). Both indigo carmine and cresyl violethave been used to detect the lack of a normal mucosal ‘net-work pattern’ and crypt openings, which correlate with in-flammatory activity and histology. Indigo carmine dyespraying was also used in 85 patients with ulcerative colitisundergoing surveillance examinations. Thirty-two werefound to have flat polyps, and 19% of these were neoplastic(24). These lesions were ill-defined before dye spraying.

Tattooing of the colon has also been described exten-sively. Tattooing is clinically applicable to mark the site of apossible malignant polyp, and to follow lesions longitudi-nally in vivo. Tattooing with India ink has been safe andwithout side effects, but systematic studies are lacking (10).Shatz and colleagues (11) followed 54 patients with Indiaink tattoos for 1.5 to 117 months. All tattoos persisted with-out symptoms or complications, and in 84 biopsies, 78 werewithout inflammation. Six biopsies demonstrated only mildinflammation; five of these were studied six months later,with three having no residual inflammation. Results fromthis and other reports of nearly 200 patients with colonic tat-toos indicate that India ink is not only durable but also safe.

CONCLUSIONSChromoscopy has many uses as an adjunctive technique dur-ing gastrointestinal endoscopy. This technique is underusedclinically, and its full clinical and research applicability is yet

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to be determined (2). Appropriately designed and imple-mentable outcomes studies will determine the place tissuestaining has in endoscopic practice.

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22. Axelrad A, Fleischer D, Geller A, et al. High-resolutionchromo-endoscopy for the diagnosis of diminutive colon polyps:implications for colon cancer screening. Gastroenterology1996;110:1253-8.

23. Matsumoto T, Kuroki F, Mizuno M, Nakamura S, Iida M. Applicationof magnifying chromoscopy for the assessment of severity in patientswith mild to moderate ulcerative colitis. Gastrointest Endosc1997;45:400-5.

24. Jaramillo E, Watanabe M, Befrits R, Ponce de Leon E, Rubio C,Slezak P. Small, flat colorectal neoplasia in long-standing ulcerativecolitis detected by high-resolution electronic video endoscopy.Gastrointest Endosc 1996;44:15-22.

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