Transanal mesorectal excision: early outcomes in Australia and New Zealand.

BACKGROUND: Transanal total mesorectal excision (taTME) aims to overcome some of the technical challenges faced when operating on mid and low rectal cancers. Specimen quality has been confirmed previously, but recent concerns have been raised about oncological safety. This multicentre prospective study aimed to evaluate the safety of taTME among early adopters in Australia and New Zealand. METHODS: Data from all consecutive patients who had taTME for rectal cancer from July 2014 to February 2020 at six tertiary referral centres in Australasia were recorded and analysed. RESULTS: A total of 308 patients of median age of 64 years underwent taTME. Some 75.6 per cent of patients were men, and the median BMI was 26.8 kg/m2. The median distance of tumour from anal verge was 7 cm. Neoadjuvant chemoradiotherapy was administered to 57.8 per cent of patients. The anastomotic leak rate was 8.1 per cent and there was no mortality within 30 days of surgery. Pathological examination found a complete mesorectum in 295 patients (95.8 per cent), a near-complete mesorectum in seven patients (2.3 per cent), and an incomplete mesorectum in six patients (1.9 per cent). The circumferential resection margin and distal resection margin was involved in nine patients (2.9 per cent), and two patients (0.6 per cent) respectively. Over a median follow-up of 22 months, the local recurrence rate was 1.9 per cent and median time to local recurrence was 30.5 months. CONCLUSION: This study showed that, with appropriate training and supervision, skilled minimally invasive rectal cancer surgeons can perform taTME with similar pathological and oncological results to open and laparoscopic surgery.

An assessment of an Australasian pathway for the introduction of transanal total mesorectal excision (taTME).

AIM: To evaluate the use of a pathway for the introduction of transanal total mesorectal excision (taTME) into Australia and New Zealand. METHOD: A pathway for surgeons with an appropriate level of specialist training and baseline skill set was initiated amongst colorectal surgeons; it includes an intensive course, a series of proctored cases and ongoing contribution to audit. Data were collected for patients who had taTME, for benign and malignant conditions, undertaken by the initial adopters of the technique. RESULTS: A total of 133 taTME procedures were performed following the introduction of a training pathway in March 2015. The indication was rectal cancer in 84% of cases. There was one technique-specific visceral injury, which occurred prior to that surgeon completing the pathway. There were no cases of postoperative mortality; morbidity occurred in 27.1%. The distal resection margin was clear in all cases of rectal cancer, and the circumferential resection margin was positive in two cases. An intact or nearly intact total mesorectal excision was obtained in more than 98% of cases. CONCLUSION: This study demonstrates the safe and controlled introduction of a new surgical technique in a defined surgeon population with the use of a pathway for training. The authors recommend a similar pathway to facilitate the introduction of taTME to colorectal surgical practice.

Systematic review of the prevalence of faecal incontinence.

BACKGROUND: Faecal incontinence (FI) is widely recognized as a significant problem in the community. Conjecture exists around the proportion of the population affected. This systematic review evaluated studies reporting the community prevalence of FI in terms of methodology, design and definitions. METHODS: MEDLINE, Embase, CINAHL, the Cochrane Collaboration and National Guideline databases were searched for studies investigating the prevalence of FI in community-based adults published from January 1966 to February 2015. Study data, including methodology, sample size, response rate, definition of FI and prevalence rates, were extracted on to a pro forma and appraised critically. Where possible, FI prevalence estimates were pooled. RESULTS: Thirty studies were analysed from 4840 screened articles. FI prevalence estimates varied from 1·4 to 19·5 per cent. This variation was explained by differences in data collection method and two factors within definitions of FI: type of stool and frequency of FI episodes. When these factors were accounted for, the FI prevalence at a threshold of at least once per month for liquid or solid stool was 8·3-8·4 per cent for face-to-face or telephone interviews, and 11·2-12·4 per cent for postal surveys. The pooled prevalence rate from studies for functional FI (defined by ROME II criteria) was 5·9 (95 per cent c.i. 5·6 to 6·3) per cent. CONCLUSION: When comparable methodologies and definitions are used, studies produce remarkably similar prevalence rates in different community populations. FI remains an unspoken symptom, with lower rates reported in personal interviews compared with anonymous postal questionnaires.

The use of local anaesthesia in haemorrhoidal banding: a randomized controlled trial.

AIM: Rubber band ligation is a common office procedure for the treatment of symptomatic haemorrhoids. It can be associated with pain and vasovagal symptoms. The effect of local anaesthetic use during banding was studied. METHOD: A single-blinded randomized controlled trial was carried out in the colorectal outpatient clinic. Patients presenting with symptomatic haemorrhoids suitable for banding were prospectively recruited and randomized to undergo the procedure with local anaesthetic or without (control). Submucosal bupivacaine was injected immediately after banding just proximal to the site. Vasovagal symptoms were assessed at the time of banding and pain scores (visual analogue scale) were recorded at the conclusion of the procedure, after 15 min, and on leaving the clinic. RESULTS: Seventy-two patients (40 local anaesthetic injection, group 1; 32 no injection, group 2) were recruited. The mean ages were 50 and 54 years respectively, the median duration of symptoms was 12 months in each group and the median number of haemorrhoids banded was three in each group. The mean pain score on leaving the clinic was 2.6 (95% CI 2.1, 3.1) in group 1 and 4.1 (95% CI 3.3, 5.0) (P = 0.04) in group 2. There were no complications related to local anaesthetic use. No significant difference in vasovagal symptoms was found (P = 0.832). CONCLUSION: Local anaesthetic injection at the time of banding is simple and safe. It may reduce patient discomfort following banding of haemorrhoids.

Determining levels of fecal incontinence in the community: a New Zealand cross-sectional study.

BACKGROUND: Fecal incontinence is a socially stigmatized condition, and its prevalence in the community has been problematic to quantify because of difficulty with its definition. OBJECTIVE: This study estimates the community prevalence of fecal incontinence in New Zealand by 3 scales of measurement: patient perceptions of a "problem with bowel control," their symptoms, and their quality of life. DESIGN/MAIN OUTCOME MEASURES: A postal survey of 2000 people, aged >18, randomly selected from the national electoral roll, was performed. This used a validated, reliability-tested, anonymous questionnaire, the Comprehensive Fecal Incontinence Questionnaire, incorporating the identification of a "problem with bowel control," the Fecal Incontinence Severity Index, and the Fecal Incontinence Quality of Life Scale. RESULTS: The response rate was 68.7%. A total of 14.7% (95% CI: 12.6-16.7) of participants "felt they had a problem with bowel control" and 12.4% (95% CI: 10.5-14.5) had fecal incontinence when defined using the Fecal Incontinence Severity Index table as "leakage of liquid or solid stool ≥ 1/month." In terms of quality of life, 26.8% of the population (95% CI: 24.2-29.4) noted some impairment on the Fecal Incontinence Quality of Life Scale. In total, 155 (13.2%) participants reported at least 2 of the 3 possible diagnostic measures, and this may provide a way to incorporate the 3 measures into a new definition of fecal incontinence. LIMITATIONS: This study incorporated a new "generic" question enquiring about an individual's perception of a bowel control problem and also introduced a "cutoff" value for Fecal Incontinence Quality of Life Scale to attempt to identify those with any impairment "due to accidental bowel leakage." CONCLUSIONS: This study helps to highlight some of the challenges involved with suitably identifying those who have fecal incontinence within the community. The prevalence rate of 13.2% represents a realistic measure of the burden of fecal incontinence in the general population, and further research in this area is recommended.

Randomized controlled trial of preoperative oral carbohydrate treatment in major abdominal surgery.

BACKGROUND: Major surgery is associated with postoperative insulin resistance which is attenuated by preoperative carbohydrate (CHO) treatment. The effect of this treatment on clinical outcome after major abdominal surgery has not been assessed in a double-blind randomized trial. METHODS: Patients undergoing elective colorectal surgery or liver resection were randomized to oral CHO or placebo drinks to be taken on the evening before surgery and 2 h before induction of anaesthesia. Primary outcomes were postoperative length of hospital stay and fatigue measured by visual analogue scale. RESULTS: Sixty-nine and 73 patients were evaluated in the CHO and placebo groups respectively. The groups were well matched with respect to surgical procedure, epidural analgesia, laparoscopic procedures, fasting period before induction and duration of surgery. Postoperative changes in fatigue score from baseline did not differ between the groups. Median (range) hospital stay was 7 (2-35) days in the CHO group and 8 (2-92) days in the placebo group (P = 0.344). For patients not receiving epidural blockade or laparoscopic surgery (20 CHO, 19 placebo), values were 7 (3-11) and 9 (2-48) days respectively (P = 0.054). CONCLUSION: Preoperative CHO treatment did not improve postoperative fatigue or length of hospital stay after major abdominal surgery. A benefit is not ruled out when epidural blockade or laparoscopic procedures are not used. REGISTRATION NUMBER: ACTRN012605000456651 (http://www.anzctr.org.au).

Systematic review on ventral rectopexy for rectal prolapse and intussusception.

OBJECTIVE: This systematic review assesses the effectiveness of ventral rectopexy (VR) surgery for treatment of rectal prolapse (RP) and rectal intussusception (RI) in adults. Method MEDLINE, EMBASE, Scopus and other relevant databases were searched to identify studies. Randomized controlled trials or nonrandomized studies with more than 10 patients receiving ventral mesh rectopexy surgery were considered for the review. RESULTS: Twelve nonrandomized case series studies with 728 patients in total are included in the review. Seven studies used the Orr-Loygue procedure (VR with posterior rectal mobilization to the pelvic floor) and five studies used VR without posterior rectal mobilization. Overall weighted mean percentage decrease in faecal incontinence (FI) rate was 45%. The weighted mean percentage decrease in constipation rate was 24%. Weighted mean recurrence rate was 3.4%. CONCLUSIONS: There are limitations in published literature on VR. The available data indicate that VR has low recurrence and improves FI in patients suffering from these conditions. There is a greater reduction in postoperative constipation if VR is used without posterior rectal mobilization.

A predictive model for local recurrence after transanal endoscopic microsurgery for rectal cancer.

BACKGROUND: The outcome of local excision of early rectal cancer using transanal endoscopic microsurgery (TEM) lacks consensus. Screening has substantially increased the early diagnosis of tumours. Patients need local treatments that are oncologically equivalent to radical surgery but safer and functionally superior. METHODS: A national database, collated prospectively from 21 regional centres, detailed TEM treatment in 487 subjects with rectal cancer. Data were used to construct a predictive model of local recurrence after TEM using semiparametric survival analyses. The model was internally validated using measures of calibration and discrimination. RESULTS: Postoperative morbidity and mortality were 14.9 and 1.4 per cent respectively. The Cox regression model predicted local recurrence with a concordance index of 0.76 using age, depth of tumour invasion, tumour diameter, presence of lymphovascular invasion, poor differentiation and conversion to radical surgery after histopathological examination of the TEM specimen. CONCLUSION: Patient selection for TEM is frequently governed by fitness for radical surgery rather than suitable tumour biology. TEM can produce long-term outcomes similar to those published for radical total mesorectal excision surgery if applied to a select group of biologically favourable tumours. Conversion to radical surgery based on adverse TEM histopathology appears safe for p T1 and p T2 lesions.

Stapled versus handsewn methods for ileocolic anastomoses.

BACKGROUND: Ileocolic anastomoses are commonly performed for right-sided colon cancer and Crohn's disease. The anastomosis may be constructed using a linear cutter stapler or by suturing. Individual trials comparing stapled versus handsewn ileocolic anastomoses have found little difference in the complication rate but they have lacked adequate power to detect potential small difference. To our knowledge, this is the first systematic review specifically investigating ileocolic anastomosis. OBJECTIVES: To compare outcomes of ileocolic anastomoses performed using stapling and handsewn techniques. The hypothesis tested was that the stapling technique is associated with fewer complications. SEARCH STRATEGY: MEDLINE, EMBASE, Cochrane Colorectal Cancer Group specialised register SR-COLOCA, Cochrane Library were searched for randomised controlled trials comparing use of a linear cuter stapler with any type of suturing technique for ileocolic anastomoses in adults from 1970 to 2005. Abstracts presented to the following society meetings between 1970 and 2002 were handsearched: American Society of Colon and Rectal Surgeons, the Association of Coloproctology of Great Britain and Ireland, European Association of Coloproctology. SELECTION CRITERIA: Randomised controlled trials comparing use of linear cutter stapler (isoperistaltic side to side or functional end to end) with any type of suturing technique in adults. DATA COLLECTION AND ANALYSIS: Eligible studies were selected and their methodological quality assessed. Relevant results were extracted and missing data sought from the authors. RevMan 4.2 Analysis version 1.0.5 was used to perform meta-analysis when there were sufficient data. Sub-group analyses for cancer and inflammatory bowel disease as indication for ileocolic anastomoses were performed. MAIN RESULTS: After obtaining individual data from authors for studies that include other anastomoses, six trials (including one unpublished) with 955 ileocolic participants (357 stapled, 598 handsewn) were included. The three largest trials had adequate allocation concealment. Stapled anastomosis was associated with significantly fewer anastomotic leaks compared with handsewn (S=5/357, HS=36/598, OR 0.34 [0.14, 0.82] p=0.02). One study performed routine radiology to detect asymptomatic leaks. For the sub-group of 825 cancer patients in four studies, stapled anastomosis led to significant fewer anastomotic leaks (S=4/300, HS=35/525, OR 0.28 [0.10, 0.75] p=0.01). There were too few Crohn's disease patients to perform sub-group analysis. All other outcomes: stricture, anastomotic haemorrhage, anastomotic time, re-operation, mortality, intra-abdominal abscess, wound infection, length of stay, showed no significant difference. AUTHORS' CONCLUSIONS: Stapled functional end to end ileocolic anastomosis is associated with fewer leaks than handsewn anastomosis.

Local excision of rectal tumours by transanal endoscopic microsurgery.

BACKGROUND: Transanal endoscopic microsurgery (TEM) allows locally complete excision of rectal tumours and provides an alternative to conventional surgery for benign tumours. However, its role in the curative treatment of invasive carcinoma is controversial. The aim of this study was to determine the morbidity and long-term results for rectal tumours excised by TEM. METHODS: Between February 1993 and January 2005, 200 patients underwent TEM for excision of adenomas (148) or carcinomas (52). The median tumour distance from the anal verge was 8 (range 1-16) cm. RESULTS: Mortality and morbidity rates were 0.5 and 14.0 per cent respectively. At a median follow-up of 33 (range 2-133) months, local recurrence had developed in 11 patients (7.6 per cent) with an adenoma. Histological examination of carcinomas revealed pathological tumour (pT) stage 1 in 31 patients, pT2 in 17 and pT3 in four. Immediate salvage surgery was performed in seven patients (13 per cent). At a median follow-up of 34 (range 1-102) months, eight patients (15 per cent) with carcinomas had developed local recurrence. The overall and disease-free 5-year survival rates for patients with carcinomas were 76 and 65 per cent respectively. CONCLUSION: TEM is an appropriate surgical treatment option for benign rectal tumours. For carcinomas, it is oncologically safe provided that resection margins are clear, but strict patient selection is required.

Randomized clinical trial of botulinum toxin plus glyceryl trinitrate vs. botulinum toxin alone for medically resistant chronic anal fissure: overall poor healing rates.

PURPOSE: This study was designed to assess whether addition of glyceryl trinitrate to botulinum toxin improves the healing rate of glyceryl trinitrate-resistant fissures over that achieved with botulinum toxin alone. METHODS: Patients were randomized between botulinum toxin plus glyceryl trinitrate (Group A) and botulinum toxin plus placebo paste (Group B). Patients were seen at baseline, four and eight weeks, and six months. The primary end point was fissure healing at eight weeks. Secondary end points were symptomatic relief, need for surgery, side effects, and reduction in maximum resting and squeeze pressures. RESULTS: Thirty patients were randomized. Two-thirds of patients had maximum anal resting pressures below or within the normal range at entry to the study. Healing rates in both treatment groups were disappointing. There was a nonsignificant trend to better outcomes in Group A compared with Group B in terms of fissure healing (47 vs. 27 percent), symptomatic improvement (87 vs. 67 percent), and resort to surgery (27 vs. 47 percent). CONCLUSIONS: There is some evidence to suggest that combining glyceryl trinitrate with botulinum toxin is superior to the use of botulinum toxin alone for glyceryl trinitrate-resistant anal fissure. The poor healing rate may reflect the fact that many of the patients did not have significant anal spasm at trial entry.

A novel model used to compare water-perfused and solid-state anorectal manometry.

BACKGROUND: Anal pressures are commonly measured using water-perfused and solid-state manometers. We constructed a dynamic model of the anus to compare the agreement and reproducibility of the two types of manometers. METHODS: The model system was constructed using a pig anorectum together with an inflatable bowel sphincter. The pig anorectum was mounted on a jig and the sphincter was inserted external to the internal sphincter. The sphincter pressure was adjusted over the range 20 to 185 mmHg. At each of 24 constant sphincter pressures, triplicate readings were carried out with both manometers. The first measurement by each method was used for the comparison. The replicate measurements were used to calculate measures of repeatability for each method. RESULTS: Measurements by the two manometers were highly correlated (r=0.97). Measurements by the solid state manometer were higher than the water-perfused manometer by 8.1+/-12.2 mmHg (mean+/-SD). Precision (coefficient of variation) for the solid-state manometer (2.8%) was better than for the water-perfused manometer (8.3%). CONCLUSIONS: The new model of the anal canal shows promise as a tool for assessing physiological interventions. The solid-state manometer has many advantages over the water-perfused manometer, providing more consistent measurements at clinically relevant pressures.

Local invasion of the bladder with colorectal cancers: surgical management and patterns of local recurrence.

PURPOSE: Colorectal cancers may be adherent to the urinary bladder. To achieve oncologic clearance of the cancer, en bloc bladder resection should be performed. This study describes the multicenter experiences of en bloc bladder resection for colorectal cancer in the major New Zealand public hospitals. METHODS: A retrospective database of patients undergoing surgery for colorectal cancer adherent to the bladder between 1984 and 1999 was constructed. Data was analyzed for age, gender, disease stage, and outcome (local recurrence and survival). RESULTS: Fifty-three patients were identified: International Union Against Cancer and American Joint Committee on Cancer Stage 1=0; Stage 2=23; Stage 3=22; Stage 4=6; unknown=2. Forty-five had en bloc partial cystectomy performed, four en bloc total cystectomy, and four had the adhesions disrupted and no bladder resection. The most common site of the primary colorectal cancer is sigmoid colon, with local invasion into the dome of the bladder. All patients who did not have en bloc resection developed local recurrence and died from their disease. Mean follow-up was 62 months. The extent of bladder resection did not seem important in determining local recurrence. CONCLUSIONS: En bloc resection of the urinary bladder should be performed if the patient is to be offered an optimal oncologic resection for adherent colorectal cancer. The decision to perform total rather than partial cystectomy should be based on the anatomic location of the tumor. Because the sigmoid is usually the primary site, most patients will not have received preoperative radiation. Therefore, postoperative radiotherapy may reduce local recurrence in these patients.

Skip metastases in colon cancer: assessment by lymph node mapping using molecular detection.

BACKGROUND: Colon cancer has been assumed to spread sequentially through the regional lymphatic bed, with skip metastases occurring in only 1% to 3% of cases. Molecular techniques allow the detection of occult metastases, but to date have not been applied to assess the pattern of regional lymphatic spread of colon cancer. METHODS: Fifty-five tumors from 54 patients with colonic adenocarcinoma were studied. Lymph node mapping was performed on fresh colonic specimens recording the position of each node on an anatomical diagram. Half of each lymph node was submitted for routine histology examination and half assayed for keratin 20 gene expression by reverse transcription-polymerase chain reaction. Logistic regression was used to analyze the distribution of histologic and occult metastases. RESULTS: A total of 1084 lymph nodes were dissected (median, 19 nodes; range, 4-52). Sixty-four lymph nodes from 20 tumors had histologically evident metastases and 76 lymph nodes from 13 tumors had occult metastases. There was no difference in the distribution of either histologic or occult metastases among paracolic, intermediate, and apical node groups. Ten patients had evidence of anatomical skip lesions after lymph node mapping and molecular analysis, only 1 of which was histologically detectable. CONCLUSIONS: This study demonstrates a higher incidence of skip metastases in colon cancer assessed by molecular techniques than has previously been reported, challenging the concept of sequential development of early lymph node metastases.

Diagnostic use of the sentinel node in colon cancer.

PURPOSE: The aim of this study was to compare the lymphatic drainage of colon cancer with the anatomic distribution of histologic and submicroscopic lymph node metastases. METHODS: Patients attending for colectomy were eligible to enter the study. At the commencement of surgery, 40 MBq of 99mTc colloidal antimony sulfide in 2 ml of Patent Blue dye was injected subserosally around the tumor. Resection was completed in a standard fashion. After resection, specimens were imaged with a gamma camera to determine the site of sentinel lymph nodes, and then dissected, recording the position of the lymph nodes on an anatomic diagram. Recovered lymph nodes were bisected, one-half for routine histology and one-half for assessment by keratin 20 (K20) reverse transcription polymerase chain reaction. The kappa measure of agreement was used to assess concordance between sentinel nodes and histologic and submicroscopic metastases. RESULTS: Four hundred fifty-six lymph nodes were dissected from 26 tumors and evaluated using lymphoscintigraphy and lymph node mapping. Sentinel nodes were evident in 23 tumors (88 percent). The sensitivity of sentinel nodes involvement as a predictor of metastatic disease was 55 percent (95 percent confidence interval, 23-83), with a false negative (nondiagnostic) rate of 45 percent. Sentinel nodes involved the apical group in four tumors, and represented anatomic "skip" lesions in four tumors. CONCLUSIONS: Direct lymphatic drainage to the apical group does occur in colon cancer; however, sentinel node mapping of colon cancer by this technique is of little clinical value because of the poor concordance between lymph node metastases and sentinel nodes.

Keratin 20 is a specific marker of submicroscopic lymph node metastases in colorectal cancer: validation by K-RAS mutations.

Lymph node status has major prognostic importance in colorectal cancer and greater precision in the diagnosis of lymph node metastases should provide better prognostic and therapeutic guidance. Keratin 20 (K20) gene expression has been used as a marker of lymph node metastases, but the evidence for this remains circumstantial. This study has therefore sought to determine K20 specificity and to correlate K20 expression with mutant K-RAS expression, in order to provide direct evidence that K20 expression in lymph nodes of colorectal cancer patients genuinely reflects metastatic disease. Specificity of K20 expression was established against a range of tissue types and 289 lymph nodes from 41 non-cancer control patients. K20 expression was restricted to gastrointestinal epithelia and was only present in one of the 289 control lymph nodes, giving a calculated specificity of 97.6% (95% confidence limits: 87.1-99.9%). Forty-two tumour samples were analysed for the presence of K-RAS codon 12 gene mutations using a RT-PCR mutant allele-specific amplification (MASA) technique. Thirteen tumours (31%) had codon 12 mutations detected by MASA and these were further analysed to determine the exact nature of the mutation. MASA was then used to screen the lymph nodes from these patients for the presence of the tumour-specific K-RAS transcript and the results were compared with K20 RT-PCR and histopathology from the same samples. Whilst K-RAS MASA was not as sensitive as K20 RT-PCR, there was substantial agreement between the assays. There were no K20-negative lymph nodes which were found to be K-RAS MASA-positive, whereas seven nodes in four patients were K20-positive and K-RAS-negative, in keeping with the differences in assay sensitivity. These results further validate K20 as a marker by providing greater certainty that what is being detected represents occult metastatic disease.

Analysis of potential markers for detection of submicroscopic lymph node metastases in breast cancer.

We have developed sensitive assays for cytokeratin (K) 8, 16, 19, stromelysin 3 (ST3), MUC1 and maspin mRNAs using reverse transcription polymerase chain reaction (RT-PCR) and used these to assess lymph node status in patients undergoing surgery for breast cancer. In addition the RT-PCR assays were tested against lymph nodes from non-cancer patients to determine their specificity. Despite high sensitivity RT-PCR assays for K8, K16, K19, ST3 and maspin were not found to be useful as markers of submicroscopic disease as transcripts of these genes were detected in the great majority of control lymph nodes tested. Expression of MUC1 was also not found to be useful as it was both insensitive and non-specific. The importance of assessing potential markers against an adequately sized control population is demonstrated, as failure to do so can lead to erroneous conclusions.

Analysis of IGF2 gene imprinting in breast and colorectal cancer by allele specific-PCR.

The insulin-like growth factor II (IGF2) gene is imprinted with the paternal allele expressed and the maternal one silent. Loss of imprinting (LOI) of IGF2 has been suggested to play a role in the development of tumours, but the reported incidence of IGF2 LOI in tumours shows considerable variation, which may stem from different methodologies employed. In particular, partial digestion of reverse transcriptase-polymerase chain reaction (RT-PCR) products by restriction enzymes can lead to inaccurate measurements. To overcome the problem of partial enzymatic digestion, a novel method termed allele specific-polymerase chain reaction (AS-PCR) has recently been reported, which provides a significant advance over enzymatic digestion. A second problem with measurements of biallelic IGF2 transcription is that the co-amplification of contaminating genomic DNA during the RT-PCR step can lead to an overestimation of the frequency of biallelic IGF2 expression. To investigate the extent of this problem, total RNA from breast and colorectal cancer was analysed using two methods. The first method involved a first-round PCR using cDNA generated with primers spanning exons 8 and 9 (exon connection), followed by a second round of AS-PCR using primers from within exon 9. The second method used only AS-PCR with primers from within exon 9. The result was that the exon-connection approach was more accurate, thereby highlighting a significant problem in imprinting analyses where genomic DNA contamination cannot be completely ruled out.