Equine grass sickness is associated with major abnormalities in the ultrastructure of skeletal neuromuscular junctions.

BACKGROUND: Equine grass sickness (EGS) is a frequently fatal multisystem neuropathy of equids. The aetiology is unknown; proposed causes include toxicoinfection with Clostridium botulinum and a mycotoxicosis. The effect of EGS on the organisation and structural integrity of the skeletal neuromuscular junction (NMJ), the target of botulinum neurotoxins (BoNTs), is unknown. OBJECTIVES: To compare the organisation and structural integrity of skeletal NMJs from EGS horses, control horses and one horse with a presumptive diagnosis of botulism. STUDY DESIGN: Blinded, retrospective case control. METHODS: NMJs in samples of diaphragm or intercostal muscle from six EGS horses, three control horses and one equine botulism case were compared using electron microscopy, morphometry and confocal light microscopy. RESULTS: A significantly higher percentage of EGS NMJs had abnormal morphology (EGS 72.2%, 95% CI 55.6-84.4; Controls 6.9%, 1.7-23.8; OR 35.1, 8.47-244.8; p < 0.001). EGS NMJs had a significantly lower mean volume fraction occupied by synaptic vesicles (SVs) (EGS 18.7%, 12.6-28.0; Controls 36.3%, 20.8-63.4; p = 0.024). EGS NMJs had evidence of accelerated SV exocytosis and SV depletion, accumulation of neurofilament-like material in terminal boutons and/or bouton degeneration. NMJs from the botulism horse had dense packing of SVs towards the presynaptic membrane active zone, consistent with BoNT intoxication, but had absence of the abnormalities identified in EGS NMJs. MAIN LIMITATIONS: Group sizes were limited by difficulties obtaining suitably processed samples. Ages of control and EGS horses differed. Botulism was diagnosed based on clinical and post mortem findings. CONCLUSIONS: EGS is associated with major changes in skeletal NMJ ultrastructure that are inconsistent with the effects of BoNTs. SV depletion may reflect increased exocytosis coupled with reduced repopulation of SVs via anterograde axonal transport and endocytosis, consistent with the action of an excitatory presynaptic toxin and/or neurotransmitter reuptake inhibitor. Skeletal NMJs represent a previously unrecognised target for the toxin that causes EGS.

Enzyme replacement therapy for mucopolysaccharidosis VI: long-term cardiac effects of galsulfase (Naglazyme®) therapy.

Characteristic cardiac valve abnormalities and left ventricular hypertrophy are present in untreated patients with mucopolysaccharidosis type VI (MPS VI). Cardiac ultrasound was performed to investigate these findings in subjects during long-term enzyme replacement therapy (ERT) with recombinant human arylsulfatase B (rhASB, rhN-acetylgalactosamine 4-sulfatase, galsulfase, Naglazyme®). Studies were conducted in 54 subjects before ERT was begun and at specific intervals for up to 96 weeks of weekly infusions of rhASB at 1 mg/kg during phase 1/2, phase 2, and phase 3 trials of rhASB. At baseline, mitral and aortic valve obstruction was present and was significantly greater in those ≥12 years of age. Mild mitral and trace aortic regurgitation were present, the former being significantly greater in those <12 years. Left ventricular hypertrophy, with averaged z-scores ranging from 1.6-1.9 SD greater than normal, was present for ages both <12 and ≥12 years. After 96 weeks of ERT, ventricular septal hypertrophy regressed in those <12 years. For those ≥12 years, septal hypertrophy was unchanged, and aortic regurgitation increased statistically but not physiologically. Obstructive gradients across mitral and aortic valves remained unchanged. The results suggest that long-term ERT is effective in reducing intraventricular septal hypertrophy and preventing progression of cardiac valve abnormalities when administered to those <12 years of age.

Thoroughbred Racehorses in Hong Kong Require Vitamin D Supplementation to Mitigate the Risk of Low Vitamin D Status.

There is a paucity of data relating to the vitamin D status of racehorses. We hypothesised that the management of racehorses in Hong Kong (HK) predisposes to low vitamin D status unless they receive dietary supplementation. Serum concentrations of 25-hydroxyvitamin D2 (25OHD2), 25-hydroxyvitamin D3 (25OHD3) and total 25-hydroxyvitamin D (total 25OHD) for 79 non-grazing HK racehorses were compared with those for 22 racehorses training in the United Kingdom (UK) that grazed for ≥1 h/d, and for which published data exists. A nested group of 41 HK horses was sampled twice to determine the effect of the duration in HK on vitamin D status. The HK horses had significantly lower serum concentrations of total 25OHD and 25OHD2 than the UK horses; 25OHD2 was undetectable in 15/79 HK sera and serum concentrations of 25OHD2 declined with the duration in HK. The main determinants of vitamin D status were assessed using linear regression; the retained variables were the 25OHD3 concentration and the duration in HK. The inverse relationship between the serum concentrations of 25OHD2 and 25OHD3, previously identified in humans, was observed for the first time in horses. In conclusion, HK racehorses have low serum 25OHD2 and total 25OHD concentrations and rely on D3 supplementation to maintain adequate vitamin D status. Further study is required to determine the optimal form of dietary vitamin D supplementation for Thoroughbred racehorses.

The effect of season, management and endocrinopathies on vitamin D status in horses.

BACKGROUND: Vitamin D deficiency is common in humans and is increasingly linked to the pathogenesis of a multitude of diseases including obesity and metabolic syndrome. The biology of vitamin D in horses is poorly described; the relative contribution of the diet and skin synthesis to circulating concentrations is unclear and associations with the endocrine disease have not been explored. OBJECTIVES: To determine the relationship between management, season and endocrine disease and vitamin D status in horses. STUDY DESIGN: Cross-sectional cohort study. METHODS: Plasma concentrations of 25-hydroxyvitamin D2 (25(OH)D2 ) and D3 (25(OH)D3 ) were measured by liquid chromatography-tandem mass spectrometry in 34 healthy unsupplemented grazing ponies and 22 stabled Thoroughbreds receiving supplementary vitamin D3 in feed. A nested group of 18 grazing ponies were sampled on long and short days (>12 and <12 h of light/day) to determine the effect of sunlight exposure. In addition, the relationships between age, sex, adiposity, serum insulin, adrenocorticotropic hormone and vitamin D status were assessed in a mixed group of 107 horses using a linear regression model. RESULTS: All animals had a measurable level of 25(OH)D2 (median 10.7 nmol/L) whilst 25(OH)D3 was only detected in Thoroughbreds receiving D3 supplementation. Thoroughbreds had lower concentrations of 25(OH)D2 than ponies (7.4 vs. 12.6 nmol/L, p < 0.01). In grazing ponies, 25(OH)D2 concentrations were significantly higher on long days compared to short days (14.4 vs. 8.7 nmol/L, p < 0.01), whilst 25(OH)D3 was undetectable. Measures of increased adiposity, but not basal insulin, were associated with higher 25(OH)D2 concentrations, conversely to humans. Increasing ACTH was associated with lower 25(OH)D2 (p < 0.01). MAIN LIMITATIONS: Vitamin D2 concentrations were not measured in grass or forage. CONCLUSIONS: In horses 25(OH)D2 is the predominant vitamin D metabolite, and there is an apparent lack of endogenous vitamin D3 production. The relationship between vitamin D and endocrine disorders in horses does not reflect that of other species and warrants further investigation.

Development and pilot-testing of a Decision Aid for use among Chinese women facing breast cancer surgery.

BACKGROUND: Women choosing breast cancer surgery encounter treatment decision-making (TDM) difficulties, which can cause psychological distress. Decision Aids (DAs) may facilitate TDM, but there are no DAs designed for Chinese populations. We developed a DA for Chinese women newly diagnosed with breast cancer, for use during the initial surgical consultation. AIMS: Conduct a pilot study to assess the DA acceptability and utility among Chinese women diagnosed with breast cancer. METHODS: Women preferred the DA in booklet format. A booklet was developed and revised and evaluated in two consecutive pilot studies (P1 and P2). On concluding their initial diagnostic consultation, 95 and 38 Chinese women newly diagnosed with breast cancer received the draft and revised draft DA booklet, respectively. Four-day post-consultation, women had questionnaires read out to them and to which they responded assessing attitudes towards the DA and their understanding of treatment options. RESULTS: The original DA was read/partially read by 66/22% (n = 84) of women, whilst the revised version was read/partially read by 74/16% (n = 35), including subliterate women (χ(2) = 0.76, P = 0.679). Knowledge scores varied with the extent the booklet was read (P1: F = 12.68, d.f. 2, P < 0.001; P2: F = 3.744, d.f. 2, P = 0.034). The revised, shorter version was graphically rich and resulted in improved perceived utility, [except for the 'treatment options' (χ(2) = 5.50, P = 0.019) and 'TDM guidance' (χ(2) = 8.19, P = 0.004) sections] without increasing anxiety (F = 0.689, P = 0.408; F = 3.45, P = 0.073). CONCLUSION: The DA was perceived as acceptable and useful for most women. The DA effectiveness is currently being evaluated using a randomized controlled trial.

Laparoscopic intragastric approach for stromal tumours located at the posterior gastric wall.

Surgical resection is indicated for gastrointestinal stromal tumour (GIST). Laparoscopic resection with clear margins is effective in gastric GIST. However, GIST located in the posterior gastric wall with close proximity to the oesophagogastric junction poses special challenges. Laparoscopic intragastric resection offers a new approach to the management of these tumours. The surgical technique, our experience and a summary of the literature are presented.

Randomized trial comparing pantoprazole infusion, bolus and no treatment on gastric pH and recurrent bleeding in peptic ulcers.

BACKGROUND: To study the effects of pantoprazole on gastric pH and recurrent bleeding after endoscopic treatment for bleeding peptic ulcers. METHODS: After endoscopic haemostasis, patients were randomly assigned to infusion group (pantoprazole 80 mg i.v. bolus followed by continuous infusion of 8 mg/h for 3 days), bolus group (pantoprazole 80 mg i.v. bolus followed by 40 mg i.v. bolus every 12 h for 3 days) and no-treatment group (no acid suppression in the first 3 days). Gastric pH was monitored. Rebleeding rate within 30 days, the need for surgery, transfusion requirement, total hospital stay, mortality rate and gastric pH were compared. RESULTS: One hundred and sixty-eight patients were included, with 15 patients excluded from the analysis. There were 54 patients in the infusion group, 49 in the bolus group and 50 in the no-treatment group. There was fewer rebleeding (3.7 vs 16.0%, P = 0.034), less operative intervention (0 vs 8.0%, P = 0.034) and shorter hospital stay (6.4 vs 8.2 days, P = 0.040) in the infusion group compared with that in no-treatment group. When the bolus group was compared with no-treatment group, there were fewer rebleed (4.1 vs 16.0%, P = 0.049) and less blood transfusion (1.5 vs 2.9 units, P = 0.007). There was no difference in mortality among the three groups. Patients who received either pantoprazole infusion or bolus had significantly higher mean pH and longer duration of pH above 6 compared with the no-treatment group. There was no difference in the rebleeding rate, transfusion requirement, need for operation and hospital stay between the infusion and bolus groups. The mean pH and the duration of pH above 6 were also similar. CONCLUSION: Pantoprazole either as infusion or bolus decreased rebleeding after endoscopic treatment for bleeding peptic ulcer.

Decellularization of placentas: establishing a protocol.

Biological biomaterials for tissue engineering purposes can be produced through tissue and/or organ decellularization. The remaining extracellular matrix (ECM) must be acellular and preserve its proteins and physical features. Placentas are organs of great interest because they are discarded after birth and present large amounts of ECM. Protocols for decellularization are tissue-specific and have not been established for canine placentas yet. This study aimed at analyzing a favorable method for decellularization of maternal and fetal portions of canine placentas. Canine placentas were subjected to ten preliminary tests to analyze the efficacy of parameters such as the type of detergents, freezing temperatures and perfusion. Two protocols were chosen for further analyses using histology, scanning electron microscopy, immunofluorescence and DNA quantification. Sodium dodecyl sulfate (SDS) was the most effective detergent for cell removal. Freezing placentas before decellularization required longer periods of incubation in different detergents. Both perfusion and immersion methods were capable of removing cells. Placentas decellularized using Protocol I (1% SDS, 5 mM EDTA, 50 mM TRIS, and 0.5% antibiotic) preserved the ECM structure better, but Protocol I was less efficient to remove cells and DNA content from the ECM than Protocol II (1% SDS, 5 mM EDTA, 0.05% trypsin, and 0.5% antibiotic).

Triaging referral to a specialist breast clinic.

BACKGROUND: The aim of this study was to determine the effectiveness of a triage system in predicting patients with malignancy among those referred to a specialist breast clinic. METHODS: A retrospective study of all referrals seen at the specialist breast clinic from January 2002 to June 2002 was conducted. The triage system allocated an urgent appointment if (i) urgent referral was requested by the referring physicians or (ii) 'non-urgent referral' was made and any one of the following 'high-risk' criteria were present: aged more than 50 years when presenting with breast lump, lump larger than 3 cm, bloody nipple discharge or physical signs suggestive of malignancy. Routine appointment was given if these conditions were not met. The outcomes of individual groups were assessed. RESULTS: Three hundred and sixty-three referrals were analysed and 44 cancers (13.2%) were diagnosed. The mean waiting time for urgent and routine appointments was 19 and 154 days, respectively. There were 108 urgent referrals and 21 (19.4%) cancers were diagnosed. Ninety-two patients were given an urgent appointment because of the presence of high-risk criteria, and 21 cancers were detected (22.8%). After the two-stage triage, breast cancer was subsequently diagnosed in only 2 out of the remaining 163 patients (1.2%) given a routine appointment. CONCLUSION: Most of the patients with cancer (96%) were given an urgent appointment through the triage system. In addition to the assessment by referring physicians, certain high-risk criteria are helpful to select patients who should be seen urgently.

The Impact of the Oncotype DX Breast Cancer Assay on Treatment Decisions for Women With Estrogen Receptor-Positive, Node-Negative Breast Carcinoma in Hong Kong.

BACKGROUND: The Oncotype DX Breast Cancer Assay is validated to assess risk of distant recurrence and likelihood of chemotherapy (CT) benefit in estrogen receptor-positive ESBC in various populations. In Hong Kong, > 80% of breast cancers are early stage breast cancer (ESBC) and > 60% of these women receive CT. This prospective study measured changes in CT type and recommendations, as well as physician impression of assay impact in a homogenous Chinese population. METHODS: Consecutive patients with estrogen receptor-positive, T1-3 N0-1mi M0 ESBC were offered enrollment. After surgery, physicians discussed treatment options with patients, then ordered the assay, then reassessed treatment recommendation considering assay results. Changes in treatment recommendation, CT utilization, physician confidence, and physician rating of influence on their treatment recommendations were measured. RESULTS: A total of 146 evaluable patients received pre- and post-testing treatment recommendations. CT recommendations (including changes in intensity of CT) were changed for 34 of 146 patients (23.3%; 95% confidence interval, 16.7%-31.0%); change in intensity occurred in 7 of 146 (4.8%). There were 27 changes in treatment recommendations of adding or removing CT altogether (18.5% change; 95% confidence interval, 12.6%-25.8%). CT recommendations decreased from 52.1% to 37.7%, a net absolute reduction of 14.4% (P < .001; 27.6% net relative reduction). Pre-assay, 96% of physicians agreed/strongly agreed that they were confident in their treatment recommendation; post-assay, 90% of physicians agreed/strongly agreed with the same statement. Thirty percent of physicians agreed/strongly agreed that the test had influenced their recommendation, similar to the proportion of changed recommendations. CONCLUSIONS: The Oncotype DX Assay appears to influence physician ESBC adjuvant treatment recommendations in Hong Kong.

Day care center attendance and diarrheal morbidity in Colombia.

This study was designed to determine whether day care center attendance was associated with increased risk of diarrheal disease among poor children in an urban, developing country setting. From July 17 to December 18, 1988, mothers of 493 Colombian children less than 5 years old (241 attendees and 252 nonattendees) were interviewed weekly about diarrheal events during the previous week. The incidence of diarrhea was greater for day care center attendees than for nonattendees (3.2 vs 2.0 episodes per child-year, P < .0005). For children less than 2 years of age, attendees experienced 7.2 episodes/child-year vs 3.5 episodes per child-year for nonattendees (P < .0005). Analyses controlling for water source and availability, excreta disposal, socioeconomic status, and duration of follow-up showed that the increased diarrheal risk was limited to children younger than 3 years of age spending more than 30 hours per week in the centers. In addition, although the risk among attendees of suffering diarrheal episodes of longer duration was fairly constant across levels of socioeconomic status, this risk was inversely proportional to socioeconomic status for nonattendees. In summary, the increase in risk of diarrhea among young, full-time day care attendees was modest, yet important, because diarrhea continues to be a major cause of morbidity and mortality in Colombian children.

Extraction, partial purification and characterization of a bacteriocin (fragicilin) produced by a strain of Bacteroides fragilis isolated from Callithrix penicillata.

A strain of Bacteroides fragilis, isolated from the marmoset Callithrix penicillata, produced protein(s) with bacteriocin activity (fragicilin). Two active fractions (36 and 150 kDa) were isolated by chromatography. The bacteriocin exhibited iso- and heteroantagonism. It remained stable between pH 3 and 10 and at 60 degrees C for 24 h. Pronase, trypsin, proteinase K and type VII protease inactivated the bacteriocin, giving evidence of its protein nature.

Microwave oven application in the extraction of fat from the subcutaneous tissue of Iberian pig ham.

The fat from subcutaneous tissue samples of Iberian pigs was extracted by solvent (2:1 chloroform-methanol) and by the action of microwave energy. The samples of liquid fat obtained by both extraction techniques were analysed by gas chromatography. Paired comparisons showed no significant differences in fatty acids in suggesting that the fat obtained by the two techniques was the same and that the extraction of fatty acids by microwaves is suitable for quality classification of Iberian pig carcasses.

The effect of areca nut on salivary and selected oral microorganisms.

The aim of this study was to investigate the effect on the growth of salivary and selected oral microorganisms of areca nut, aqueous extracts of the nut, its major alkaloid arecoline and the components tannic acid and catechin of its tannin fraction. The antibacterial properties of the above were tested on Streptococcus mutans, Streptococcus salivarius, Candida albicans and Fusobacterium nucleatum and, as a control, Staphylococcus aureus. This was followed by investigating its effect on salivary organisms cultured from the saliva after chewing boiled areca nut. Extracts inhibited the growth of the selected organisms in a concentration dependent manner, baked and boiled nuts being significantly more potent than raw nut. Growth of C. albicans was the least affected by the nut extracts. Tannic acid was strongly antibacterial but not catechin or arecoline. No antibacterial effect could be demonstrated on salivary organisms after chewing the nut for 5 minutes but exposure of saliva to the cud for 1 hour caused a significant depression of bacterial growth. It is concluded that the hydrolysable tannins in the tannin fraction, which include tannic acid, are responsible for the antibacterial properties of the nut and that prolonged intraoral exposure to the nut can suppress bacteria in the mouth.

[Anna Justina Ferreira Nery: framework in the history of Brazilian nursing]. / Anna Justina Ferreira Nery: um marco na história da enfermagem brasileira.

The object of this study is Anna Neri (1814-1880), an anonymous woman who participated in the Paraguay War (1865-1870). This is a social-historical biographical study guided by the principles of the New History. The objective of this research is to characterize and raise relevant aspects of Anna Neri's life. It is known that this woman left her domestic life to play an important role in the public space. Her motivation was genuinely affectionate, and she didn't expect to be called a nurse. Anna Neri met the requirements that would raise the social and moral status of the XXth century nursing workers, and became a symbol of Brazilian nursing.

Bone marrow mononuclear cells versus mesenchymal stem cells from adipose tissue on bone healing in an Old World primate: can this be extrapolated to humans? / Células mononucleares de medula óssea versus células-tronco mesenquimais de tecido adiposo em cicatrização de lesão óssea em primata: pode ser extrapolado para humanos?

In veterinary medicine, the cell therapy is still unexplored and there are many unanswered questions that researchers tend to extrapolate to humans in an attempt to treat certain injuries. Investigating this subject in nonhuman primates turns out to be an unparalleled opportunity to better understand the dynamics of stem cells against some diseases. Thus, we aimed to compare the efficiency of bone marrow mononuclear cells (BMMCs) and mesenchymal stem cells (MSCs) from adipose tissue of Chlorocebus aethiops in induced bone injury. Ten animals were used, male adults subjected, to bone injury the iliac crests. The MSCs were isolated by and cultured. In an autologous manner, the BMMCs were infused in the right iliac crest, and MSCs from adipose tissue in the left iliac crest. After 4.8 months, the right iliac crests fully reconstructed, while left iliac crest continued to have obvious bone defects for up to 5.8 months after cell infusion. The best option for treatment of injuries with bone tissue loss in old world primates is to use autologous MSCs from adipose tissue, suggesting we can extrapolate the results to humans, since there is phylogenetic proximity between species.(AU)

Na medicina veterinária, a terapia celular ainda é inexplorada e há muitas perguntas não respondidas, o que leva os pesquisadores a uma tendência a estender a terapia para os seres humanos, na tentativa de tratar certas lesões. Investigar esse assunto em primatas não humanos revela-se uma oportunidade sem precedentes para compreender melhor a dinâmica das células-tronco contra algumas doenças. Assim, objetivou-se comparar a eficiência das células mononucleares de medula óssea (BMMCs) e das células-tronco mesenquimais (MSCs) do tecido adiposo de Chlorocebus aetiops na lesão óssea induzida. Foram utilizados 10 animais, adultos do sexo masculino, submetidos à lesão óssea nas cristas ilíacas. As MSCs foram isoladas e cultivadas; de forma autóloga, as BMMCs foram infundidas na crista ilíaca direita e as MSCs de tecido adiposo na crista ilíaca esquerda. Após 4,8 meses, a crista ilíaca direita foi totalmente reconstruída, enquanto a crista ilíaca esquerda continuou apresentando defeito ósseo evidente por até 5,8 meses após a infusão. A melhor opção para o tratamento de lesões com perda de tecido ósseo em primatas do Velho Mundo é a utilização de MSCs autólogas de tecido adiposo, sugerindo que se podem estender os resultados para seres humanos, uma vez que há proximidade filogenética entre as espécies.(AU)

Decellularization of placentas: establishing a protocol

Biological biomaterials for tissue engineering purposes can be produced through tissue and/or organ decellularization. The remaining extracellular matrix (ECM) must be acellular and preserve its proteins and physical features. Placentas are organs of great interest because they are discarded after birth and present large amounts of ECM. Protocols for decellularization are tissue-specific and have not been established for canine placentas yet. This study aimed at analyzing a favorable method for decellularization of maternal and fetal portions of canine placentas. Canine placentas were subjected to ten preliminary tests to analyze the efficacy of parameters such as the type of detergents, freezing temperatures and perfusion. Two protocols were chosen for further analyses using histology, scanning electron microscopy, immunofluorescence and DNA quantification. Sodium dodecyl sulfate (SDS) was the most effective detergent for cell removal. Freezing placentas before decellularization required longer periods of incubation in different detergents. Both perfusion and immersion methods were capable of removing cells. Placentas decellularized using Protocol I (1% SDS, 5 mM EDTA, 50 mM TRIS, and 0.5% antibiotic) preserved the ECM structure better, but Protocol I was less efficient to remove cells and DNA content from the ECM than Protocol II (1% SDS, 5 mM EDTA, 0.05% trypsin, and 0.5% antibiotic).

Intra-operative assessment of axillary sentinel lymph nodes by frozen section-an observational study of 260 procedures.

BACKGROUND: Sentinel lymph node biopsy (SLNB) has become the standard treatment for patients with invasive breast cancer. Intra-operative frozen section allows for the detection of nodal metastases, thereby allowing a simultaneous axillary lymph node dissection (ALND) for those patients with nodal metastases. We herein report the accuracy of frozen section in the detection of SLN metastases. METHODS: Patients with operable breast cancer and clinically negative axillae were recruited for SLNB. The SLNs were identified by blue dye, an isotope, or a combination. Enlarged lymph nodes that were not SLNs were also excised. All nodes were examined by intra-operative frozen section. ALND was performed if frozen section was positive. For those without metastases on frozen section, ALND was not performed. All lymph nodes underwent further paraffin sectioning with immuno-histochemical staining. RESULTS: A total of 260 SLNB procedures were performed for invasive carcinoma over a 3-year period. The SLN was identified in 93.5% of patients. Of the 243 successful procedures, 53 had nodal metastases on frozen section. A total of 33 patients had false-negative frozen sections (false-negative rate, 38.4%), and 97% of them were less than 2 mm in size. The false-negative rate for macro-metastases, micro-metastases, and isolated tumour cells_were 2.4%, 57.7%, and 94.4%, respectively (p < 0.0001). A total of 22 patients had delayed ALND, and the re-operation rate was 8.5%. CONCLUSION: Frozen section was useful for the detection of nodal metastases in the SLNs and allowed for ALND to be performed in the same operation. The main failure of frozen sections was in the detection of micro-metastases.