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Aims: The cell block technique for assessing endoscopic ultrasound-guided fine needle aspiration (EUS-FNA) specimens from pancreatic mucinous cystic lesions (MCLs) was systematically evaluated for the first time, including comparisons with three traditional methods of assessing such specimens., Methods: The prospective arm comprised EUS-FNA specimens from EUS-suspected pancreatic MCLs. The retrospective arm comprised EUS-FNA specimens from pancreatic MCLs surgically resected before the study start. For each specimen, these data points were collected: macroscopic likelihood of mucin, cyst fluid carcinoembryonic antigen (CEA) level and presence of mucin in air-dried, direct smears and in cell block preparations., Results: The prospective and retrospective arms of the study comprised 80 and 30 EUS-FNA specimens, respectively. Seven prospective cases led to surgical resections during the study, and therefore, 37 EUS-FNA specimens were confirmed to have originated from MCLs. In the prospective arm, macroscopic mucin was suspected, cyst fluid CEA level exceeded 192 ng/mL, mucin was detected in direct smears and cell block preparations in 78%, 30%, 39% and 73% of cases, respectively. Of the 37 specimens confirmed to originate from MCLs, macroscopic mucin assessment, cyst fluid CEA level, direct smear mucin assessment and cell block mucin assessment had sensitivities for diagnosing MCL of 87%, 45%, 45% and 81%, respectively., Conclusions: Cell block preparations are as likely to identify mucin from pancreatic MCLs as macroscopic assessment but are twice as likely to diagnose MCL than direct smears and fluid CEA biochemistry. The cell block technique is easy for sample collection and processing especially because these are identical for solid and cystic pancreatic lesions., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2020. No commercial re-use. See rights and permissions. Published by BMJ.)

Mucins are high molecular weight glycoproteins with complex oligosaccharide side chains attached to the apomucin protein backbone by O-glycosidic linkage; they are found in crude mucus gels that protect epithelial surfaces in the major tracts of the body and as transmembrane proteins expressed on the apical cell surface of glandular and ductal epithelia of various organs. Changes in the sequence of glycosylation of mucins in different settings generate a variety of epitopes in the oligosaccharide side chains of mucins, including newly expressed blood-group antigens, distinguishing between normal and diseased states. Tumour-associated epitopes on mucins and their antigenicity make them suitable as immunotargets on malignant epithelial cells and their secretions, creating a surge of interest in mucins as diagnostic and prognostic markers for various diseases, and even influencing the design of mucin-based vaccines. This review discusses the emerging roles of mucins such as MUC1 and MUC4 in cancer and some other diseases, and stresses how underglycosylated and truncated mucins are exploited as markers of disease and to monitor widespread metastasis, making them useful in patient management. Furthermore the type, pattern and amount of mucin secreted in some tissues have been considered in the classification and terminology of neoplasia and in specific organs such as the pancreas. These factors have been instrumental in pathological classification, diagnosis and prognostication of neoplasia.

Breast lesions with mucin represent a broad spectrum of entities, ranging from benign fibrocystic changes with luminal mucin to mucocele-like lesions (MLL), which can be associated with banal epithelial alterations, atypical ductal hyperplasia or ductal carcinoma in situ. Occasionally invasive mucinous carcinoma can be identified in contiguity with MLL. Diagnostic challenges are enumerated, histological differentials are discussed, and a practical approach towards resolving some of these issues is provided. In addition to these lesions with abundant extracellular mucin, there are also conditions that feature stromal mucinous or myxoid material, as well as rare entities that demonstrate both epithelial extracellular and stromal mucin.

Mucinous metaplasia of the genital area is a rare condition characterised by the emergence of mucin containing cells in stratified squamous epithelium. This report describes a unique case of benign mucinous metaplasia of the vulva associated with lichen sclerosus in a 60 year old woman. Histopathology revealed cervical type metaplasia with otherwise typical lichen sclerosus. This report discusses the pathogenesis and differential diagnosis of mucinous epithelium.

Background: Patients with Barrett's oesophagus (BO) are at risk of oesophageal adenocarcinoma. Because the pattern of mucosal mucins changes during neoplastic progression, it may serve as a marker of intraepithelial neoplasia., Aims: To determine the expression pattern of mucins in neoplastic BO epithelium (high grade dysplasia) and correlate it with the expression of apoptosis markers Bax and Bcl-2., Methods: Thirty seven patients with BO were studied: 16 without intraepithelial neoplasia, six with high grade intraepithelial neoplasia (HGN), and 15 with infiltrating adenocarcinoma. Biopsies were obtained from squamous epithelium, Barrett's epithelium, and (when present) foci of suspected HGN or adenocarcinoma. MUC1-4, MUC5AC, MUC5B, MUC6, Bax, and Bcl-2 mRNA were determined by semiquantitative RT-PCR. MUC2, MUC5AC, and MUC6 protein was determined by immunoblotting., Results: Mucin expression varied between neoplastic progression stages in BO. Mucin mRNA levels were low in squamous epithelium, except for MUC4, and were at least four times higher in BO and HGN (p<0.001), but less so in adenocarcinoma. MUC4 expression was significantly lower in BO than in normal squamous epithelium, whereas in HGN and adenocarcinoma, levels were significantly higher than in BO (p = 0.037). The Bax:Bcl-2 ratio was increased in HGN compared with BO (p = 0.04). MUC2, MUC5AC, and MUC6 protein values correlated with mRNA data., Conclusions: Mucin expression varies during the development of oesophageal adenocarcinoma in BO. MUC4 could serve as a tumour marker in this process. In contrast to animal studies, upregulation of MUC4 in HGN is associated with increased apoptosis, suggesting that MUC4 plays a minor role in apoptosis regulation in BO.

Background: In Barrett's oesophagus (BO), squamous epithelium is replaced by specialised intestinal epithelium (SIE). Transcription factors associated with intestinal differentiation, such as CDX2, may be involved in BO development., Aim: To investigate CDX2 expression in BO, squamous epithelium, and oesophageal adenocarcinoma (ADC)., Methods: CDX2 expression was assessed in 245 samples-167 biopsies of the columnar lined segment and 38 squamous epithelial biopsies of 39 patients with histologically confirmed BO (10 with ADC). Forty biopsies from 20 patients with reflux oesophagitis (RO) without BO were also evaluated. CDX2 protein was investigated immunohistochemically in 138 biopsies from 16 patients with BO, four with ADC, and 20 with RO. Cdx2 and Muc2 mRNA were detected semiquantitatively using 88 BO biopsies and squamous epithelium from 19 BO patients, and when present from ADC., Results: SIE was present in 53/79 biopsies from the columnar lined segment; CDX2 protein was seen in all epithelial cells, but not in biopsies containing only gastric metaplastic epithelium (26/79), or in squamous epithelium (0/40) of patients with RO. Cdx2 mRNA was detected in all biopsies with goblet cell specific Muc2 transcription-indicative of SIE. Low Cdx2 mRNA expression was seen in 6/19 squamous epithelium samples taken 5 cm above the squamocolumnar junction of BO patients., Conclusion: CDX2 protein/mRNA is strongly associated with oesophageal SIE. Cdx2 mRNA was present in the normal appearing squamous epithelium of one third of BO patients, and may precede morphological changes seen in BO. Therefore, pathways that induce Cdx2 transcription in squamous epithelial cells may be important in BO development.

Background: Pleomorphic adenoma of the salivary gland (PA) is essentially a benign neoplasm. However, patients with recurrent PA are difficult to manage. There are rare reports on useful immunohistochemical markers to detect a high risk of recurrence when the primary lesions are resected., Aims: To find a new marker to predict the recurrence of PA., Methods: Primary lesions of PA were collected from nine patients showing subsequent recurrence and from 40 patients without recurrence during at least 10 years of follow up of the disease. Paraffin wax embedded tumour samples of the two groups were examined for the expression profiles of MUC1 (differentially glycosylated forms), MUC2, MUC4, MUC5AC, and MUC6 using immunohistochemistry. Several clinicopathological factors were also examined., Results: In univariate analysis of the factors examined, MUC1/DF3 high expression (more than 30% of the neoplastic cells stained) in the primary lesions was seen more frequently in patients with recurrence (four of nine) than in those without recurrence (three of 40; p = 0.011). Larger tumour size (more than 3.0 cm) of the primary PA was also a significant (p = 0.035) risk factor for the recurrence of PA. In multivariate analysis, only high expression of MUC1/DF3 was found to be a significant independent risk factor for the recurrence of PA (p = 0.021)., Conclusions: Expression of MUC1/DF3 in PA is a useful marker to predict its recurrence. Those patients with PA showing positive MUC1/DF3 expression should be followed up carefully.

Aims: To determine gastric expression of trefoil family factor 2 (TFF2) and MUC6 in Helicobacter pylori positive and negative subjects, and its association with antralisation at the gastric incisura., Methods: Gastric biopsies from the antrum, incisura, and body of 76 dyspeptic patients without ulcers were used for the determination of H. pylori infection, histological changes, and epithelial TFF2 and MUC6 expression., Results: In the foveola, the rates of TFF2 and MUC6 immunostaining were greater in H. pylori infected (n = 27) than in uninfected patients (n = 49) at the antrum (59.3% v 4.1% for TFF2 and 63.0% v 4.1% for MUC6; both p < 0.001) and incisura (44.4% v 2.0% for TFF2 and 48.1% v 0% for MUC6; both p < 0.001). In the deeper glands, the rates were also greater in H. pylori infected than in uninfected patients at the incisura (85.2% v 22.4% for both TFF2 and MUC6; p < 0.001). Antral-type mucosa was present at the incisura in 28 of the 76 patients. TFF2 and MUC6 expression in the foveola and deeper glands was significantly associated with antral-type mucosa, independent of H. pylori status., Conclusions: Helicobacter pylori infection increases the expression of TFF2 and MUC6 in the gastric epithelium. Aberrant TFF2 and MUC6 expression is associated with antralisation of gastric incisura.

MUCs are glycoproteins with various roles in homeostasis and carcinogenesis. Among other actions, MUC1 may inhibit cell-cell and cell-stroma interactions and function as a signal transducer, participating in cancer progression. In contrast, MUC2 is normally found only in goblet cells, where it contributes to the protective barrier function of these cells. Recently, a tumour suppressor role has been demonstrated for MUC2, and both MUC1 and MUC2 appear to have important roles in pancreatic neoplasia. MUC1 appears to be a marker of aggressive phenotype and may facilitate the vascular spread of carcinoma cells. In contrast, MUC2 is rarely detectable in aggressive pancreatic tumours, but is commonly expressed in intraductal papillary mucinous neoplasms (IPMNs), which are rare, indolent tumours, in intestinal IPMNs, and in indolent colloid carcinomas. MUC2 appears to be not only a marker of this indolent pathway, but also partly responsible for its less aggressive nature. Thus, in pancreatic neoplasia, MUC1 and MUC2 have potential diagnostic and prognostic value as markers of aggressive and indolent phenotypes, respectively, and have potential as therapeutic targets.

Background: Premalignant Barrett's oesophagus (BO) and gastric intestinal metaplasia (IM) show phenotypic variability. Incompletely differentiated sulfomucin rich gastric IM (type III) may have increased malignant potential. The types of sulfated oligosaccharide structures present in IM, BO, and colon have not been fully characterised., Aims: To compare sulfo-Lewis(a) epitope tissue distribution with high iron diamine (HID) positive sulfomucin in metaplastic, dysplastic, and neoplastic tissues from oesophagus and stomach., Methods: Sections containing gastric IM or BO (some associated with dysplasia or adenocarcinoma) were stained by the HID/alcian blue (AB) method and immunohistochemically (antibody 91.9H) to detect sulfo-Lewis(a). Based on HID/AB staining, IM was subtyped into type I (complete) or types II and III (incomplete)., Results: In total, 125 sections from 38 subjects were studied. Normal squamous oesophagus, normal gastric epithelium, and type I IM were negative for sulfomucin and sulfo-Lewis(a). In type II IM, occasional goblet cells were HID and sulfo-Lewis(a) positive, but sialomucin secreting (AB positive) columnar cells were sulfo-Lewis(a) negative. Type III IM was always sulfo-Lewis(a) positive. Sulfomucin staining in dysplasia and cancer was variable, but HID positive areas were always sulfo-Lewis(a) positive., Conclusions: Sulfo-Le(a), which is expressed on colonic mucin, is invariably present on sulfomucins in gastric IM and BO. Its presence in incomplete variants of IM and its absence from type I IM emphasises the phenotypic differences between complete and incomplete forms of metaplasia. 91.9H immunostaining is useful in IM subtyping. Characterising the molecular basis of sulfo-Lewis(a) expression may help understand the process of aberrant differentiation.

Aims: To compare the lineage specific distribution of MUC1 and MUC2 mucins in normal colorectal mucosa and adenocarcinoma and to identify pathological correlations., Methods: Paraffin wax sections from 51 colorectal cancers were examined for the expression of MUC1 and MUC2, non-O-acetyl sialic acid and the carbohydrate epitopes Lex, Ley, sialosyl-Lex, sialosyl-Tn, and Tn using standard histochemical methods., Results: MUC1, Lex and Ley co-localised with columnar cell secretions, whereas MUC2, mild periodic acid Schiff and sialosyl-Tn co-localised with goblet cell mucin in both normal and malignant tissues. Sialosyl-Lex and Tn were associated with both lineages. In normal tissues MUC1, Lex and Ley showed only trace expression by crypt base columnar cells. Cancers could be classified into four phenotypes (MUC2+/MUC1-, MUC2+/MUC1+, MUC2-/MUC1+, MUC2-/MUC1-). Particular phenotypes showed significant correlations with cancer type, lymph node spread and peritumoral lymphocytic infiltration and trends falling short of significance in relation to grade of differentiation and contiguous adenoma., Conclusions: Classification of colorectal cancer by means of lineage specific function may be relevant to both pathogenesis and prognosis.

Aims: To examine the expression of the MUC2 epithelial mucin in breast carcinoma; to relate this to patient survival., Methods: Sections from 210 breast carcinomas were stained with the anti-MUC2 core protein monoclonal antibody, 4F1, using an immunoperoxidase technique. The proportion of tumour cells positively stained and the localisation and intensity of any staining were recorded. Expression of MUC2 was compared with histological type and grade, tumour size, presence of nodal metastases, presence of oestrogen receptors, and menopausal status. The prognostic value of MUC2 expression was examined using Kaplan-Meier survival analysis., Results: MUC2 mucin was detected in 19% of cases of invasive carcinoma, in 11% of cases of carcinoma in situ, where present, but very rarely in adjacent normal breast epithelium. Presence of MUC2 was significantly associated with a shorter disease free interval (p < 0.05), although the observed difference in duration of overall survival was not significant., Conclusions: The MUC2 detected in breast carcinoma may be underglycosylated or staining may represent detection of the protein core before the completion of glycosylation. The virtual absence of 4F1 reactivity in normal breast epithelium suggests that, unlike the MUC1 mucin, the MUC2 mucin is not highly expressed by these cells. The mechanism by which expression of MUC2 affects the biology of breast tumours is unclear, although expression may be a reflection of general derepression of genes during tumour progression.

Aims: To produce and characterise a monoclonal antibody specific for O-acetylated sialomucin and to assess its use in immunohistochemistry on a panel of normal and diseased intestinal tissue samples., Methods: Mouse monoclonal antibodies were developed following immunisation with highly purified human colonic mucin. One of these (MMM-17) showed strong binding to mucin throughout the normal colon with relative lack of binding to colon cancer tissue. The binding epitope of MMM-17 was then characterised by screening for agglutination activity against a panel of human and animal erythrocytes and by assessment of its binding to a range of normal and chemically treated slot blotted mucins. Further immunohistochemical studies were then performed on formalin fixed, normal, and diseased human intestinal samples., Results: Binding of MMM-17 to slot blotted human colonic mucin was reduced by 38 (SD 14%) (n = 4) by alkali treatment of the mucin, sequential alkali and sialidase treatment completely abolished binding. Sialidase treatment alone, however, caused only an 11 (11%) reduction in binding. MMM-17 failed to agglutinate any human, rabbit, rat or mouse erythrocytes. These findings were compatible with specificity of MMM-17 for sialomucins O-acetylated at the C-7 or C-8 positions on the sialic acid. Strong staining by MMM-17 was found in all goblet cells throughout all 40 normal colonic and rectal samples studied, but staining was absent in seven of 13 colorectal carcinomas. Normal duodenum (n = 16) and normal ileum (n = 3) all showed occasional positive goblet cells. The normal gastric antral mucosa was generally negative B MMM-17, but in all of 15 cases of gastritis with intestinal metaplasia the metaplastic glands were strongly positive for MMM-17., Conclusion: Monoclonal antibody MMM-17 has specificity for O-acetylated sialomucins and its binding depends both on the position of O-acetylation and on the adjacent oligosaccharide structure. Preliminary studies using the antibody on archival tissue samples support the previous reports of reduced O-acetylation in colon cancer demonstrated by indirect histochemistry and show the neo-formation of O-acetylated sialomucin in intestinal metaplasia in the stomach.

Aims: To determine whether acidic mucin staining, lectin histochemistry using Wisteria floribunda agglutinin, and immunohistochemistry using the monoclonal antibody EAB 903 are of benefit in distinguishing between hyperplastic and carcinomatous prostatic glandular tissue in needle biopsy specimens., Methods: Formalin fixed, paraffin wax embedded prostatic needle biopsy specimens of benign and malignant tissue were examined. Alcian blue-periodic acid Schiff staining was performed on 33 benign and 34 malignant cases. Wisteria floribunda agglutinin (WFA) binding sites were demonstrated by the avidin-biotin peroxidase (ABC) technique with and without neuraminidase pretreatment on 34 benign cases and 32 malignant cases. EAB903 anticytokeratin antibody binding sites were demonstrated using both an indirect immunoperoxidase (IIP) technique and an avidin-biotin peroxidase complex method on seven benign and 31 malignant cases., Results: Acidic mucin staining was found in 17 of 34 malignant glands and was weakly positive in five of 33 benign glands. WFA positivity before neuraminidase pretreatment was present in 29 of 32 malignant glands and in 19 of 34 benign glands. After neuraminidase all benign and malignant cases showed positivity. EAB 903 positivity was seen in 11 of 31 malignant glands using the IIP technique and in two of 31 malignant glands using the ABC technique. In seven benign cases there was positivity in all glands using the IIP method with predominant basal cell staining in three and superficial cell staining in four. In benign cases using the ABC method two cases were negative., Conclusions: None of the three methods studied showed sufficient sensitivity and specificity to allow their recommendation for routine diagnostic use.

Peroxidase conjugated lectins were used to analyse the glycoproteins of small intestinal mucins in normal infants and those with cystic fibrosis to ascertain whether there are any detectable histochemical differences in saccharide composition. A significant decrease in Lotus tetragonolobus (LTG) binding fucose was shown in normal small intestinal mucin starting around 36 weeks' gestation with total absence of staining at term and beyond. In contrast, the age matched patients with cystic fibrosis showed persistent and intense LTG binding of fucose. These results provide the first clear histochemical evidence that cystic fibrosis mucin is abnormal and confirm the findings of previous biochemical studies.

A method for the simultaneous demonstration of lysozyme and mucins in 39 cases of gastric adenomas differentiated two intermediate cell types. The first was similar to a columnar cell comprising a single cell population which covered extensive areas of the adenomas. This cell type often showed supranuclear lysozyme reactivity and apical neutral mucins, sialomucins, and sulphomucins in variable amounts. The second cell type was found in 11 adenomas, located mainly in the fundal area. It seemed to be a transitional form between the goblet cell and the Paneth cell. This cell type was scattered among columnar cells, occasional Paneth-like cells, and small goblet cells. These two types of intermediate cells may be regarded as abnormally differentiated integral elements of gastric adenomas. They may be associated with the neck stem cells in the cytogenesis of gastric adenomas.

A total of 1412 consecutive cases of endoscopic gastric biopsy, carried out over a four year period, were reviewed and specimens were examined histochemically to determine the prevalence of intestinal metaplasia and its variants. Three types were characterised: complete intestinal metaplasia and two classes of incomplete intestinal metaplasia (type IIa and type IIb) depending on the absence or presence, respectively, of sulphomucins within mucin secreting columnar cells. Type IIb intestinal metaplasia was significantly more common in patients with gastric carcinoma (p less than 0.001) and in those with dysplasia (p less than 0.001) than in patients with benign gastric pathology. No such association was found with either type I or type IIa intestinal metaplasia. In addition to those present in the columnar cells of type IIb intestinal metaplasia, sulphomucins were also commonly found in goblet cells of all three types of metaplasia. The presence of sulphomucins in goblet cells, however, was not significantly associated with gastric carcinoma or dysplasia. The significance of the different types of intestinal metaplasia in relation to the pathological findings is discussed.

Over a four-year period 21 cases of solitary ulcer syndrome (SUS) were studied for their clinical, histological, and mucus secretion patterns and compared with histological and mucus secretion patterns of 78 cases of non-specific proctitis collected over the same period. Normal mucus composition was found in non-specific proctitis while abnormalities of mucins with predominance of sialomucins were associated with SUS. Although histology remains the most important investigation in the diagnosis of SUS, mucin changes provide valuable additional evidence.

Forty primary carcinomas of the thyroid of different histological types were reviewed and studied histochemically, with the aim of identifying and assessing "mucin secretion". The patterns of extracellular "pure alcianophilia" and "mixed alcianophilia" were noted in 7.5% and about 50% of these tumours, respectively. A critical review of the pitfalls in methods and interpretation of mucin histochemistry--as performed in previously reported cases of "mucin secreting" or "mucinous" primary thyroid tumours--is presented. The apparent "mucin secretion" described in these unusual neoplasms could be due to histochemical staining of carbohydrate components or breakdown products of thyroglobulin and colloid.

Of 534 resected colorectal adenocarcinomas, 165 (31%) contained some mucinous components; these represented the main part of the tumour in 67 (13%). Of the mucin containing tumours, 63 (38%) were in the right colon compared with 50 (13%) of the non-mucinous ones (p less than 0.001). Patients with predominantly mucinous tumours were significantly older than those with non-mucinous tumours, and they tended to present with tumours at a more advanced stage. A multivariate analysis did not show any significant independent prognostic influence of the mucinous component except when this had a predominantly signet ring cell pattern.

Although epithelial mucin has been demonstrated in ordinary mammary duct carcinomas, it is formed profusely in the uncommon mucous carcinomas. A histochemical study of mucous carcinomas of the breast and colon showed that both types produce similar mucins, viz, acid and neutral, although the mammary carcinomas produce more neutral mucin and small amounts of sulphated mucins are produced by colonic tumours. Similar studies on sections of normal breast and cystic mammary dysplasia have shown that intracellular neutral mucin was present in the ducts and lobules, and that intracellular acid mucin was present within cells of the terminal ducts and lobules. It was demonstrated enzymatically that the acid mucins produced by the normal and neoplastic tissues is sialic acid. Hence it can be shown that the mucin produced by mucous carcinomas is of epithelial rather than connective tissue type. It is contended that mucous carcinoma of breast arises by metaplasia of duct cells that have already undergone malignant change and not from the lobules of the breast.

Epithelial mucins of the normal terminal ileum, caecum, colon, and rectum of man are unique in that they alone exhibit staining following the periodate-borohydride/KOH/PAS technique. Application of this technique enables one to differentiate those mucin-producting metastases arising from adenocarcinoma of the lower gastrointestinal tract from those arising elsewhere, and may occasionally be useful in determining the site of the primary tumour when it is in doubt. Furthermore, it was found to be especially useful in distinguishing between primary adenocarcinoma of the lung and metastases in the lung from adenocarcinoma of the lower gastrointestinal tract.

Ninety two tissue blocks from the left colon and 52 from the right colon were obtained from 112 patients with familial adenomatous polyposis (FAP). Tissues from 137 patients with other conditions served as controls. Within the main study a smaller investigation was performed to compare sections from the left and right colon in the same subject. Several well known histochemical techniques were used to investigate possible changes in sulphation, sialic acid structure (loss of O-acetyl substituents), and changes in the ratio of sialic acid to neutral sugars. In patients with FAP, as in controls, there was increased expression of periodic acid Schiff positive mucin and fucose in the right colon. The only difference between patients with FAP and controls was the indirect demonstration of less neutral mucin in the right colon in FAP, but this did not seem to affect neutral sugars binding to UEA-1, PNL, or HPA. As in the general population, a small proportion of patients with FAP showed a lack of O-acetyl substituted sialic acid. Sialic acid heterogeneity probably has a genetic basis, but this is not associated with the genetic defect underlying FAP.

It has been suggested that diffuse changes in epithelial mucins may help to identify a subgroup of patients with ulcerative colitis who are at increased risk of developing colorectal cancer. Colonoscopic biopsy specimens were taken from 11 colitic patients shortly before colectomies were performed, in which cancer was identified. These patients were matched to colitic patients without cancer for age, sex, and duration of disease. Sections were stained with high iron diamine and alcian blue (HID-AB), mild periodic acid Schiff, and for peanut lectin binding sugar sequences. The sections were assessed blindly and graded semiquantitatively. Most showed ulcerative colitis in remission or mild active disease. There were no differences between the study and control groups. Metaplastic foci were more common in the group who had cancer. These showed increased staining of mucus by mild periodic acid Schiff and increased binding by peanut lectin to the supranuclear (Golgi) zone. Epithelial dysplasia (detected in four of the patients with cancer showed focal intense binding by peanut lectin to cytoplasm, mucus, and glycocalyx. Although dysplasia differed histochemically from normal tissue, special techniques did not facilitate its diagnosis.

Characteristic morphological and histochemical changes in mucosa adjacent to colorectal adenocarcinomas have been described. It has been suggested that this type of mucosa, labelled transitional mucosa (TM) because of its specific association with colorectal adenocarcinoma, is indicative of a premalignant change. In an investigation of mucosa adjacent to anal tumours extending into the rectum and mucosa from solitary ulcer syndrome and colostomies the mucosal alterations described in TM could be demonstrated. Thus TM is not specifically related to colorectal adenocarcinoma and probably arises as a secondary phenomenon. The claim that TM represents a premalignant change cannot be substantiated.

A simple histochemical procedure for assessing relative amounts of neutral and acidic sugars in mucin glycoproteins, and its application in the study of cyclical changes of human cervical mucins, is described. This procedure, the saponification/selective periodate oxidation/borohydride reduction/alcian blue pH 2.5/periodic acid Schiff (KOH/PA*/Bh/Ab 2.5/PAS) method, uses a selective oxidation step to remove the PAS positivity of sialic acid; thus only neutral sugars stain positively with PAS, and acidic sugars (O-sulphate esters and carboxyl groups) stain with alcian blue. This differs from the KOH/Ab/PAS technique which stains sialic acid residues with both alcian blue and PAS. Applying the KOH/PA*/Bh/Ab 2.5/PAS technique to the study of cyclical changes of human cervical mucins, a decreased neutral:acidic sugar ratio in the secretory phase mucins compared with those of the proliferative phase was found. This difference was not seen with KOH/Ab/PAS staining in the same cases. The techniques and reagents used in this procedure can be easily applied in a clinical histopathology laboratory.

A comparison of routine and special histochemical methods that were applied to the epithelial mucins of small intestine from patients with Crohn's disease and from normal controls showed that the normal small intestine (ileum) goblet cells secrete a predominantly non-sulphated sialomucin and that, in contradistinction to the colon, the neuraminidase insensitivity of the sialic acids of the small intestine was not due to either O-acylation at C4 or an ester substituent at C1. Presumably this implies that the protection against enzyme attack afforded to the mucosa by the mucin coat in the small intestine utilises a different mechanism from that in the colon and that, although in many respects the small intestinal mucins in Crohn's disease, and in normal controls are similar, there is an increase in side-chain O-acylated sialic acids in such mucins in Crohn's disease. This difference has not been described before, probably because it can be seen only after staining such sections by the PAT/KOH/PAS and the PBT/KOH/PAS techniques.

The periodic acid-thionin Schiff/potassium hydroxide/periodic acid-Schiff (PAT/KOH/PAS) procedure has been used to investigate the histochemical staining characteristics of the mucins found in adenocarcinoma and villous lesions of the large intestine. The 46 blocks examined represented 58 lesions from 37 patients, all of whom had had resections for carcinoma of the colon. tin sharp contrast to normal colon, none of the adenocarcinomas stained red with the PAT/KOH/PAS. With two exceptions the poorly and moderately differentiated adenocarcinomas stained blue, whereas of the well differentiated lesions half were blue and half purple. The malignant villous lesions demonstrated the same trends, although a larger percentage were purple. None of the benign lesions stained blue. It is suggested that malignancy in the colon is accompanied by an increase in blue staining in the PAT/KOH/PAS technique and that such staining may be of value in the interpretation of highly atypical adenoma where it might identify the onset of malignancy. This change in staining indicates a distinct alteration in the chemistry of the mucins which we interpret as a reduction in the degree of side chain O-acylation of their constituent sialic acids.

Orcein-alcian blue staining, a new method for the simultaneous demonstration of sulphated and sialomucins in gastrointestinal epithelium was compared with the standard high iron diamine-alcian blue technique. Sections were oxidised with potassium permanganate and decolourised in oxalic acid. They were stained with orcein for four hours, differentiated for a few seconds in acid alcohol, and then counterstained with alcian blue for half to one minute. There was a good correlation of results between the two methods. Orcein-alcian blue is a safer, cheaper, and quicker method than high iron diamine-alcian blue which can be safely introduced into routine laboratories for the study of acid mucins in the gastrointestinal diseases.

Fifteen surgical specimens from patients with carcinoma of the colon and rectum were studied. Scrapings from normal mucosa distant from the tumour and from macroscopically normal mucosa adjacent to the tumour (;transitional') were used for chemical estimation of hexosamines, sialic acid, and proteins. The presence of hexosamines and sialic acid was confirmed in both normal and transitional mucosa. Transitional mucosa showed increased levels of total hexosamines and sialic acid as compared with the normal and this was accompanied by an increase in neuraminidase-sensitive sialic acids. The present data have been compared with previous histochemical and autoradiographic studies and it is suggested that the changes described in the transitional mucosa are transformations representing an early stage of carcinogenesis.

Tumours of the vermiform appendix were studied. The majority proved to be carcinoid tumours of the usual argentaffin-positive type but a substantial minority, generally tumours of minute size, were found to have a somewhat different histological pattern and were argentaffinnegative. Some contained argyrophil granules. The non-argentaffin tumours are considered to form a subgroup of carcinoid tumours.

One hundred and twenty-one rectal biopsies from 99 patients with carcinoma of the rectum or lower sigmoid colon were investigated using a high iron-diamine-Alcian blue technique for sulphated and non-sulphated acid mucins. It was found that in normal rectal mucosa sulphomucins are the main carbohydrate component of the goblet cell mucin. This normal ;mucous pattern' changes in the ;transitional' mucosa (histological normal mucosa adjacent to carcinoma) where there is an increase of non-sulphated acid mucins concomitantly with a decrease or absence of sulphated groups in 60 to 90% of the cases according to the type of tumour. The same type of changes in mucus were observed in the ;transitional' mucosa surrounding adenomatous polyps and papillary adenomas; they were not marked in areas around carcinoma in situ and not observed in the metaplastic polyps. These changes seem to be in direct relationship to the grade of differentiation and invasiveness of the tumour. The histochemical changes in the mucins seem to be in favour of a malignant potential in the so-called neoplastic polyps. The high iron diamine-Alcian blue, because of its ;specificity', consistent results, and easy technique is recommended for routine use together with haematoxylin and eosin staining in the diagnosis of premalignant changes.