Virtual Staining of Nonfixed Tissue Histology.

Surgical pathology workflow involves multiple labor-intensive steps, such as tissue removal, fixation, embedding, sectioning, staining, and microscopic examination. This process is time-consuming and costly and requires skilled technicians. In certain clinical scenarios, such as intraoperative consultations, there is a need for faster histologic evaluation to provide real-time surgical guidance. Currently, frozen section techniques involving hematoxylin and eosin (H&E) staining are used for intraoperative pathology consultations. However, these techniques have limitations, including a turnaround time of 20 to 30 minutes, staining artifacts, and potential tissue loss, negatively impacting accurate diagnosis. To address these challenges, researchers are exploring alternative optical imaging modalities for rapid microscopic tissue imaging. These modalities differ in optical characteristics, tissue preparation requirements, imaging equipment, and output image quality and format. Some of these imaging methods have been combined with computational algorithms to generate H&E-like images, which could greatly facilitate their adoption by pathologists. Here, we provide a comprehensive, organ-specific review of the latest advancements in emerging imaging modalities applied to nonfixed human tissue. We focused on studies that generated H&E-like images evaluated by pathologists. By presenting up-to-date research progress and clinical utility, this review serves as a valuable resource for scholars and clinicians, covering some of the major technical developments in this rapidly evolving field. It also offers insights into the potential benefits and drawbacks of alternative imaging modalities and their implications for improving patient care.

Intraoperative margin assessment during radical prostatectomy: is microscopy frozen in time or ready for digital defrost?

Intraoperative frozen section (IFS) is used with the intention to improve functional and oncological outcomes for patients undergoing radical prostatectomy (RP). High resource requirements of IFS techniques such as NeuroSAFE may preclude widespread adoption, even if there are benefits to patients. Recent advances in fresh-tissue microscopic digital imaging technologies may offer an attractive alternative, and there is a growing body of evidence regarding these technologies. In this narrative review, we discuss some of the familiar limitations of IFS and compare these to the attractive counterpoints of modern digital imaging technologies such as the speed and ease of image generation, the locality of equipment within (or near) the operating room, the ability to maintain tissue integrity, and digital transfer of images. Confocal laser microscopy (CLM) is the modality most frequently reported in the literature for margin assessment during RP. We discuss several imitations and obstacles to widespread dissemination of digital imaging technologies. Among these, we consider how the 'en-face' margin perspective will challenge urologists and pathologists to understand afresh the meaning of positive margin significance. As a part of this, discussions on how to describe, categorize, react to, and evaluate these technologies are needed to improve patient outcomes. Limitations of this review include its narrative structure and that the evidence base in this field is relatively immature but developing at pace.

Impact of frozen section on long-term outcomes in robot-assisted laparoscopic prostatectomy.

OBJECTIVES: To compare 1-year functional and 5-year oncological outcomes of men undergoing robot-assisted laparoscopic prostatectomy (RALP) with neurovascular structure-adjacent frozen-section examination (NeuroSAFE) with those in men undergoing RALP without NeuroSAFE (standard of care [SOC]). SUBJECTS AND METHODS: Men undergoing RALP in our centre between 1 January 2009 and 30 June 2018 were enrolled from a prospectively maintained database. Patients were excluded if they had undergone preoperative therapy or postoperative adjuvant therapy or were enrolled in clinical trials. Patients were grouped based on use of NeuroSAFE. Follow-up was censored at 5 years. The primary outcome was difference in time to biochemical recurrence (BCR) on multivariable analysis, defined as prostate-specific antigen (PSA) >0.2 ng/L on two consecutive measurements. Secondary outcomes were difference in 1-year erectile dysfunction and incontinence. RESULTS: In the enrolment period, 1199 consecutive men underwent RALP, of whom 1140 were eligible, including 317 with NeuroSAFE and 823 with SOC. The median PSA follow-up was 60 months in both groups. Rates of 5-year BCR were similar on Kaplan-Meier survival curve analysis (11% vs 11%; P = 0.9), as was time to BCR on multivariable Cox proportional hazards modelling (hazard ratio 1.2; P = 0.6). Compared with the SOC group at 1 year, the NeuroSAFE group had similar unadjusted rates of incontinence (5.1% vs 7.7%) and lower unadjusted impotence (57% vs 80%). On multivariable analysis, NeuroSAFE patients had equivalent risk of incontinence (odds ratio [OR] 0.59, 95% CI 0.17-1.6; P = 0.4) but significantly reduced risk of erectile dysfunction (OR 0.37, 95% CI 0.22-0.60; P < 0.001). CONCLUSIONS: For men undergoing RALP, compared with SOC, NeuroSAFE patients had equivalent time to BCR and risk of 1-year incontinence, and significantly lower risk of 1-year erectile dysfunction.

Evaluation of margins during radical prostatectomy: confocal microscopy vs frozen section analysis.

OBJECTIVES: To test the performance of ex vivo fluorescence confocal microscopy (FCM; Vivascope 2500M-G4), as compared to intra-operative frozen section (IFS) analysis, to evaluate surgical margins during robot-assisted radical prostatectomy (RARP), with final pathology as the reference standard. METHODS: Overall, 54 margins in 45 patients treated with RARP were analysed with: (1) ex vivo FCM; (2) IFS analysis; and (3) final pathology. FCM margins were evaluated by two different pathologists (experienced [M.I.: 10 years] vs highly experienced [G.R.: >30 years]) as strongly negative, probably negative, doubtful, probably positive, or strongly positive. First, inter-observer agreement (Cohen's κ) between pathologists was tested. Second, we reported the sensitivity, specificity, positive predictive (PPV) and negative predictive value (NPV) of ex vivo FCM. Finally, agreement between ex vivo FCM and IFS analysis (Cohen's κ) was reported. For all analyses, four combinations of FCM results were evaluated. RESULTS: At ex vivo FCM, the inter-observer agreement between pathologists ranged from moderate (κ = 0.74) to almost perfect (κ = 0.90), according to the four categories of results. Indeed, at ex vivo FCM, the highly experienced pathologist reached the best balance between sensitivity (70.5%) specificity (91.8%), PPV (80.0%) and NPV (87.1%). Conversely, on IFS analysis, the sensitivity, specificity, PPV and NPV were, respectively, 88.2% vs 100% vs 100% vs 94.8%. The agreement between the ex vivo FCM and IFS analyses ranged from moderate (κ = 0.62) to strong (κ = 0.86), according to the four categories of results. CONCLUSION: Evaluation of prostate margins at ex vivo FCM appears to be feasible and reliable. The agreement between readers encourages its widespread use in daily practice. Nevertheless, as of today, the performance of FCM seems to be sub-par when compared to the established standard of care (IFS analysis).

Reconsidering the Role of Frozen Section in Sentinel Lymph Node Biopsy for Mastectomy Patients.

INTRODUCTION: Axillary lymph node dissection was recommended for mastectomy patients with more than two nodal metastases from sentinel lymph node biopsy. Conventionally, intraoperative frozen section was sent routinely to reduce the need for second-stage axillary lymph node dissection; however, recent global trend has seen decreasing usage of the intraoperative analyses. This pilot study conducted in Thailand aimed to evaluate the role of intraoperative frozen section of sentinel lymph node biopsy in early-stage breast cancer patients who underwent mastectomy. METHODS: A 5-y retrospective study of 1773 patients was conducted in Thailand. The inclusion criteria were early-stage breast cancer patients with either radiologically negative nodes, or radiographically borderline nodes found to be negative on fine needle aspiration who underwent mastectomy and sentinel lymph node biopsy. Reoperations were indicated when three or more nodal metastases were detected on the pathological analysis. The reoperation rate prevented by frozen section and the reoperation rate needed for those with permanent section alone were reported. RESULTS: Among 265 patients, 202 patients underwent concomitant intraoperative frozen section while the remaining 63 patients underwent permanent section alone. Six patients (3.0%) from the frozen section group and one patient (1.6%) from the permanent section group were found with more than two nodal metastases. Despite using intraoperative frozen sections, only one patient from each group required reoperation. There was no significant difference in the number of patients requiring reoperation between the frozen section group and the permanent section group. CONCLUSIONS: Our study provides strong evidence to all surgeons that in early breast cancer patients undergoing mastectomy, sentinel lymph node biopsy with permanent section analysis alone may not lower the standard of care compared to using additional intraoperative frozen section analysis. Adopting this practice may lead to decreased operation costs, operative time, and anesthetic side effects.

The utility of intraoperative marrow margin frozen section in extremity bone sarcoma resection.

BACKGROUND AND OBJECTIVES: Intraoperative frozen section analysis is commonly used to evaluate marrow margins during extremity bone sarcoma resections, but its efficacy in the era of magnetic resonance imaging is debated. This study aimed to compare the accuracy of intraoperative frozen section assessment with final pathology, assess its correlation with gross intraoperative margin assessment, and evaluate its impact on surgical decision making. METHODS: Consecutive patients undergoing extremity bone sarcoma resections from 2010 to 2022 at a single sarcoma center were included. Intraoperative frozen section and gross margin assessments were compared to final pathology using positive predictive values (PPV) and negative predictive values (NPV). Changes in surgical decisions due to positive intraoperative margins were recorded. RESULTS: Of 166 intraoperative frozen section marrow margins, four were indeterminant/positive, with two false positive/indeterminant findings and two false negatives compared to final pathology. Gross intraoperative assessment had no false positives and two false negatives. Frozen section analysis yielded a PPV of 50% (95% confidence interval [CI]: 16%-84%) and NPV of 98.8% (95% CI: 97%-100%), while gross assessment had a PPV of 100% (95% CI: 16%-100%) and NPV of 98.8% (95% CI: 97%-100%). Positive frozen section margins led to additional resections in three of four cases. CONCLUSIONS: Intraoperative frozen section analysis did not offer added clinical value beyond gross margin assessment in extremity bone sarcoma resections. It exhibited a low PPV and led to unnecessary additional resections. Gross intraoperative assessment proved adequate for margin evaluation, potentially saving time and resources.

Does the use of intraoperative frozen section of bone marrow from the cut end of the mandible help assess the adequacy of bone margins following mandibulectomy for oral cancer?

BACKGROUND: The adequacy of the cut end of the mandible following a segmental mandibulectomy done for oral cancer intraoperatively is at times assessed using a frozen section (FS) of the bone marrow (BM) at the cut ends. The study aimed to assess its utility to guide the intraoperative decision on the adequacy of bony margins. MATERIALS AND METHODS: All patients with oral squamous cell carcinoma (OSCC) who underwent segmental mandibulectomy from January 2012 to December 2021 at our institute and for whom intraoperative FS of BM was utilized were included. We analyzed the sensitivity, specificity, and positive and negative predictive value (PPV, NPV) of this in predicting positive bone margins. RESULTS: A total of 457 patients were included in the study. The majority of the cases were per premium cases (n = 372, 81.4%). The median age of the cohort was 52 years (range: 22-80 years). Most patients had T4 disease (n = 406, 88.8%). On FS, BM was positive in only 18 patients (3.9%) for whom the bone margin was revised. BM biopsy report in the final histopathology was positive in 12 patients (2.2%). The sensitivity, specificity PPV, and NPV were 52.3%, 98.65%, 64.7%, and 97.7% respectively. No factors predicting BM positivity on FS could be identified in this cohort. CONCLUSIONS: The BM FS was positive in only a small percentage of patients, and it helped in reducing the bone margin positivity rate from 3.9% to 2.2% only. Hence the intraoperative BM FS seems to have limited utility as seen from our study.

Multiplex Intraoperative Rapid Immunohistochemistry with Noncontact Antibody Mixing for Distinguishing the Histologic Phenotype of Lung Cancer.

INTRODUCTION: Determining a surgical strategy for early-stage lung cancer requires an accurate histologic diagnosis. Immunohistochemistry (IHC) enables reliable diagnosis of histological types but requires more time and more tumor tissue slides than hematoxylin and eosin staining. We aimed to assess the clinical validity of a new rapid multiplex IHC technique utilizing alternating current (AC) mixing for intraoperative lung cancer diagnosis. METHODS: Forty-three patients who underwent radical resection of lung cancers were enrolled in a retrospective observational study. Frozen sections were prepared from lung tumor samples, and rapid IHC employing AC mixing was implemented alongside a multiplex IHC protocol targeting thyroid transcription factor-1 + cytokeratin 5, desmoglein 3 + Napsin A, and p63 + tripartite motif containing 29. We then evaluated the concordance between intraoperative diagnoses derived from rapid multiplex IHC and final pathology. RESULTS: The concordance rate between the pathological diagnosis made with added rapid multiplex IHC and the final pathology was 93.0% (Cohen's 𝜅 coefficient = 0.860 and 95% CI: 0.727-0.993). When considering only adenocarcinoma and squamous cell carcinoma, the diagnoses were in agreement for all cases. CONCLUSIONS: We suggest rapid multiplex IHC as a promising tool for determining surgical strategies for lung tumors.

Low-Grade Adenosquamous Carcinoma of the Breast Masquerading as a Fibroepithelial Lesion on Core Biopsy: A Challenging Case.

INTRODUCTION: Diagnosing low-grade adenosquamous carcinoma (LGASC) presents significant challenges due to its subtle morphology, variable immunohistochemical expression, and resemblance to benign lesions like radial scar and complex sclerosing lesions. CASE PRESENTATION: We present a case of a 53-year-old woman with a subareolar mass initially thought to be a fibroepithelial neoplasm on core biopsy. Subsequent wide excision revealed LGASC with oestrogen receptor expression (weak to moderate intensity, 40% of tumour cells). CONCLUSION: These findings, rarely reported, highlight the difficulty of diagnosing LGASC on small tissue samples.

Feasibility of intraoperative pathologic examination for sentinel lymph nodes during sentinel node navigation surgery in early gastric cancer: results of pathologic protocol for SENORITA trial.

BACKGROUND: During sentinel node navigation surgery in patients with gastric cancer, intraoperative pathologic examination of sentinel nodes is crucial in determining the extent of surgery. In this study, we evaluated the feasibility and accuracy of intraoperative pathologic protocols using data from a prospective, multicenter, randomized trial. METHODS: A retrospective analysis was conducted using data from the SEntinel Node ORIented Tailored Approach trials from 2013 to 2016. All sentinel lymph nodes were evaluated during surgery with hematoxylin-eosin (HE) staining using a representative section at the largest plane for lymph nodes. For permanent histologic evaluation, sentinel basin nodes were stained with HE and cytokeratin immunohistochemistry in formalin-fixed, paraffin-embedded (FFPE) sections and examined with HE for three deeper-step sections at 200-µm intervals. The failure rate of identification by frozen section and the metastasis rate in non-sentinel basins were investigated. RESULTS: Of the 237 patients who underwent sentinel node basin dissection, 30 had lymph node metastases on permanent pathology. Thirteen patients had macrometastasis confirmed in frozen sections as well as FFPE sections (failure rate: 0%). Patients with negative sentinel nodes in frozen sections but micrometastasis in FFPE sections had no lymph node recurrence during the follow-up period (0%, 0/6). However, in cases with tumor-positive nodes in frozen sections, metastases in non-sentinel basins were detected in the paraffin blocks (8.3%, 2/24). CONCLUSIONS: The single-section HE staining method is sufficient for detecting macrometastasis via intraoperative pathological examination. If a negative frozen-section result is confirmed, sentinel basin dissection can be performed safely. Otherwise, standard surgery is required.

Development and validation of a model for predicting upstage in minimally invasive lung adenocarcinoma in Chinese people.

BACKGROUND: Sublobar resection for ground-glass opacity became a recommend surgery choice supported by the JCOG0804/JCOG0802/JCOG1211 results. Sublobar resection includes segmentectomy and wedge resection, wedge resection is suitable for non-invasive lesions, but in clinical practice, when pathologists are uncertain about the intraoperative frozen diagnosis of invasive lesions, difficulty in choosing the appropriate operation occurs. The purpose of this study was to analyze how to select invasive lesions with clinic-pathological characters. METHODS: A retrospective study was conducted on 134 cases of pulmonary nodules diagnosed with minimally invasive adenocarcinoma by intraoperative freezing examination. The patients were divided into two groups according to intraoperative frozen results: the minimally invasive adenocarcinoma group and the at least minimally invasive adenocarcinoma group. A variety of clinical features were collected. Chi-square tests and multiple regression logistic analysis were used to screen out independent risk factors related to pathological upstage, and then ROC curves were established. In addition, an independent validation set included 1164 cases was collected. RESULTS: Independent risk factors related to pathological upstage were CT value, maximum tumor diameter, and frozen result of AL-MIA. The AUC of diagnostic mode was 71.1% [95%CI: 60.8-81.3%]. The independent validation included 1164 patients, 417 (35.8%) patients had paraffin-based pathology of invasive adenocarcinoma. The AUC of diagnostic mode was 75.7% [95%CI: 72.9-78.4%]. CONCLUSIONS: The intraoperative frozen diagnosis was AL-MIA, maximum tumor diameter larger than 15 mm and CT value is more than - 450Hu, highly suggesting that the lung GGO was invasive adenocarcinoma which represent a higher risk to recurrence. For these patients, sublobectomy would be insufficient, lobectomy or complementary treatment is encouraged.

Intraoperative pathologic diagnosis of central nervous system lymphomas: A comparison of frozen and permanent section diagnoses, and the significance of preoperative imaging.

BACKGROUND: Central nervous system (CNS) lymphomas, either primary or secondary in origin, are rare malignant tumors affecting the brain, spinal cord, or leptomeninges. Diagnosis of CNS lymphomas is complicated by their diverse clinical presentations, radiological features, and histopathological characteristics. Although frozen section (FS) analysis is commonly employed for various CNS tumors, its role and accuracy in CNS lymphoma diagnosis are less explored. In this study, we conducted a comparative analysis to assess the impact of knowledge of preoperative imaging on enhancing the accuracy of FS diagnosis in CNS lymphomas. METHODS: Data collection involved a retrospective review of CNS lymphoma patients from January 2009 to August 2021. Patients who underwent intraoperative consultation were included, excluding those with prior cortisone treatment. The dataset incorporated patient demographics, classification as primary or secondary lymphoma, radiological preliminary diagnoses, FS diagnosis, and permanent section diagnosis. We employed various archived materials, including FSs, touch imprint slides, crush cytology slides, H&E-stained sections, and immunohistochemical stains, and re-evaluated all slides for diagnostic validation. RESULTS: Our study included 25 patients, of whom 60 % were female and had a mean age of 56.5 years. Preoperative radiology data were available for 80 % of cases, with preliminary diagnoses commonly including lymphoma and/or metastasis. Intraoperative consultation results indicated lymphoma in 18 (72 %) patients, with discordance observed in 28 % of cases when compared to permanent section diagnoses. Most permanent section diagnoses were diffuse large B-cell lymphomas (92 %), with the remainder being T-cell non-Hodgkin lymphoma (4 %) and follicular lymphoma (4 %). Intraoperative misdiagnoses were significantly associated with the absence of knowledge of preoperative imaging. CONCLUSION: Our study demonstrates the reliability of FS diagnosis for CNS lymphomas during surgery, with a favorable complete concordance rate of 72 % when compared to permanent diagnoses. Importantly, lack of knowledge of preoperative imaging significantly impaired diagnostic accuracy in FS, emphasizing the need for close collaboration between pathologists and radiologists.

Prediction of lymph node metastasis in patients with papillary thyroid cancer based on radiomics analysis and intraoperative frozen section analysis: A retrospective study.

INTRODUCTION: To evaluate the diagnostic efficiency among the clinical model, the radiomics model and the nomogram that combined radiomics features, frozen section (FS) analysis and clinical characteristics for the prediction of lymph node (LN) metastasis in patients with papillary thyroid cancer (PTC). METHODS: A total of 208 patients were randomly divided into two groups randomly with a proportion of 7:3 for the training groups (n = 146) and the validation groups (n = 62). The Least Absolute Shrinkage and Selection Operator (LASSO) regression was used for the selection of radiomics features extracted from ultrasound (US) images. Univariate and multivariate logistic analyses were used to select predictors associated with the status of LN. The clinical model, radiomics model and nomogram were subsequently established by logistic regression machine learning. The area under the curve (AUC), sensitivity and specificity were used to evaluate the diagnostic performance of the different models. The Delong test was used to compare the AUC of the three models. RESULTS: Multivariate analysis indicated that age, size group, Adler grade, ACR score and the psammoma body group were independent predictors of lymph node metastasis (LNM). The results showed that in both the training and validation groups, the nomogram showed better performance than the clinical model, albeit not statistically significant (p > .05), and significantly outperformed the radiomics model (p < .05). However, the nomogram exhibits a slight improvement in sensitivity that could reduce the incidence of false negatives. CONCLUSION: We propose that the nomogram holds substantial promise as an effective tool for predicting LNM in patients with PTC.

Challenging pitfalls in frozen section pathology: a case of mandible ghost cell odontogenic carcinoma and the literature review.

BACKGROUND: Ghost cell odontogenic carcinoma (GCOC) is a rare malignancy characterized by the presence of ghost cells, preferably in the maxilla. Only slightly more than 50 case reports of GCOC have been documented to date. Due to the rarity of this tumor and its nonspecific clinical criteria, there is a heightened risk of misdiagnosis in clinical examination, imaging findings, and pathology interpretation. CASE PRESENTATION: A 50-year-old male patient presented to the hospital due to experiencing pain in his lower front teeth while eating for the past 2 months. Upon examination, a red, hard, painless mass was found in his left lower jaw, measuring approximately 4.0 cm × 3.5 cm. Based on the malignant histological morphology of the tumor and the abundant red-stained keratinized material, the preoperative frozen section pathology misdiagnosed it as squamous cell carcinoma (SCC). The surgical resection specimen pathology via paraffin section revealed that the tumor was characterized by round-like epithelial islands within the fibrous interstitium, accompanied by a large number of ghost cells and some dysplastic dentin with infiltrative growth. The malignant components displayed marked heterogeneity and mitotic activity. Additionally, a calcified cystic tumor component of odontogenic origin was observed. Hemorrhage, necrosis, and calcifications were present, with a foreign body reaction around ghost cells. Immunoreactivity for ß-catenin showed strong nuclear positivity in tumor cells, while immunostaining was completely negative for p53. The Ki67 proliferation index was approximately 30-40%. The tumor cells exhibited diffuse CK5/6, p63, and p40 immunoreactivity, with varying immunopositivity for EMA. Furthermore, no BRAFV600E mutation was identified by ARMS-PCR. The final pathology confirmed that the tumor was a mandible GCOC. CONCLUSION: We have reported and summarized for the first time the specific manifestations of GCOC in frozen section pathology and possible pitfalls in misdiagnosis. We also reviewed and summarized the etiology, pathological features, molecular characteristics, differential diagnosis, imaging features, and current main treatment options for GCOC. Due to its rarity, the diagnosis and treatment of this disease still face certain challenges. A correct understanding of the pathological morphology of GCOC, distinguishing the ghost cells and the secondary stromal reaction around them, is crucial for reducing misdiagnosis rates.

Intraoperative rapid immunohistochemistry with noncontact antibody mixing for undiagnosed pulmonary tumors.

Knowledge of the histologic type and primary origin of pulmonary tumors is essential when preparing a surgical strategy. Intraoperative diagnosis of hematoxylin and eosin (H&E)-stained frozen sections is the gold standard, but reliable pathology requires time-consuming immunohistochemistry (IHC) to distinguish among histological types/organ origins and to analyze molecular status. The aim of this study was to evaluate the clinical reliability of a new rapid-IHC technique for intraoperative diagnosis of pulmonary tumors. In total, 169 patients with undiagnosed pulmonary tumors were enrolled in a multicenter prospective observational study. At three institutes, pulmonary tumor samples were collected through core needle biopsy and/or surgery to determine surgical strategies. Using a new device for rapid IHC, we applied a high-voltage, low-frequency alternating current (AC) field, which mixes the available antibody as the voltage is switched on/off. Rapid IHC can provide tumor histologic type/origin diagnoses within 20 min, as opposed to the 3-6 h required for conventional IHC. No false diagnoses of malignancy were rendered in any of the cases when using simple H&E staining. With H&E staining alone, the overall definitive diagnosis rate, the rate of defined tumor origin, and the rate of determined histological type were 76.92%, 85.80%, and 90.53%, respectively. When rapid IHC was added, those rates were significantly improved to 88.76%, 94.67%, and 91.72%, respectively. By providing prompt and accurate intraoperative histological/molecular analysis, rapid IHC driven by AC mixing could serve as an effective clinical tool guiding the surgical strategy for undiagnosed pulmonary tumors.

Intraoperative assessment of axillary sentinel lymph nodes by telepathology.

PURPOSE: Although axillary dissection is no longer indicated for many breast cancer patients with 1-2 positive axillary sentinel lymph nodes (ASLN), intraoperative ASLN assessment is still performed in many institutions for patients undergoing mastectomy or neoadjuvant therapy. With recent advancements in digital pathology, pathologists increasingly evaluate ASLN via remote telepathology. We aimed to compare the performance characteristics of remote telepathology and conventional on-site intraoperative ASLN assessment. METHODS: Data from ASLN evaluation for breast cancer patients performed at two sites between April 2021 and October 2022 was collated. Remote telepathology consultation was conducted via the Aperio eSlideManager system. RESULTS: A total of 385 patients were identified during the study period (83 telepathology, 302 on-site evaluations). Although not statistically significant (P = 0.20), the overall discrepancy rate between intraoperative and final diagnoses was slightly higher at 9.6% (8/83) for telepathology compared with 5.3% (16/302) for on-site assessment. Further comparison of performance characteristics of ASLN assessment between telepathology and conventional on-site evaluation revealed no statistically significant differences between deferral rates, discrepancy rates, interpretive or sampling errors, major or minor disagreements, false negative or false positive results as well as clinical impact and turn-around time (P ≥ 0.12). CONCLUSION: ASLN assessment via telepathology is not significantly different from conventional on-site evaluation, although it shows a slightly higher overall discrepancy rate between intraoperative and final diagnoses (9.6% vs. 5.3%). Further studies are warranted to ensure accuracy of ASLN assessment via telepathology.

Clinical Validation of Stimulated Raman Histology for Rapid Intraoperative Diagnosis of Central Nervous System Tumors.

Stimulated Raman histology (SRH) is an ex vivo optical imaging method that enables microscopic examination of fresh tissue intraoperatively. The conventional intraoperative method uses frozen section analysis, which is labor and time intensive, introduces artifacts that limit diagnostic accuracy, and consumes tissue. SRH imaging allows rapid microscopic imaging of fresh tissue, avoids tissue loss, and enables remote telepathology review. This improves access to expert neuropathology consultation in both low- and high-resource practices. We clinically validated SRH by performing a blinded, retrospective two-arm telepathology study to clinically validate SRH for telepathology at our institution. Using surgical specimens from 47 subjects, we generated a data set composed of 47 SRH images and 47 matched whole slide images (WSIs) of formalin-fixed, paraffin-embedded tissue stained with hematoxylin and eosin, with associated intraoperative clinicoradiologic information and structured diagnostic questions. We compared diagnostic concordance between WSI and SRH-rendered diagnoses. Also, we compared the 1-year median turnaround time (TAT) of intraoperative conventional neuropathology frozen sections with prospectively rendered SRH-telepathology TAT. All SRH images were of sufficient quality for diagnostic review. A review of SRH images showed high accuracy in distinguishing glial from nonglial tumors (96.5% SRH vs 98% WSIs) and predicting final diagnosis (85.9% SRH vs 93.1% WSIs). SRH-based diagnosis and WSI-permanent section diagnosis had high concordance (κ = 0.76). The median TAT for prospectively SRH-rendered diagnosis was 3.7 minutes, approximately 10-fold shorter than the median frozen section TAT (31 minutes). The SRH-imaging procedure did not affect ancillary studies. SRH generates diagnostic virtual histologic images with accuracy comparable to conventional hematoxylin and eosin-based methods in a rapid manner. Our study represents the largest and most rigorous clinical validation of SRH to date. It supports the feasibility of implementing SRH as a rapid method for intraoperative diagnosis complementary to conventional pathology laboratory methods.

Diagnostic accuracy of sentinel lymph node frozen section analysis in patients with early-stage cervical cancer: A systematic review and meta-analysis.

OBJECTIVE: To assess the diagnostic accuracy of intraoperative SLN frozen section analysis compared with ultrastaging in patients with early-stage cervical cancer. METHODS: A systematic literature review was conducted following the PRISMA checklist. MEDLINE (via Ovid), Embase, and the Cochrane Central Register of Controlled Trials were searched from inception until February 2023. The inclusion criteria were patients with early-stage cervical cancer (2018 FIGO stage I-II), consisting of the histological subtype squamous cell carcinoma, adenocarcinoma, or adenosquamous carcinoma (≥90% of the patients in each study), who underwent SLN detection (with any tracer) and intraoperative frozen section followed by SLN ultrastaging. Randomized controlled trials, prospective and retrospective observational studies were considered. The detection rates and measures of diagnostic accuracy were pooled using a random effects univariate model. A preplanned subgroup meta-analysis was conducted, with isolated tumor cells excluded as positive lymph nodes. The review was registered in PROSPERO (CRD42023397147). RESULTS: The search identified 190 articles, with 153 studies considered potentially eligible after removing duplicates. Fourteen studies met the selection criteria, including a total of 1720 patients. Seven studies were retrospective, and the other seven were prospective. Frozen section analysis detected 159 of 292 (54.5%) patients with lymph node metastases. In 281 patients the type of volume metastasis was reported: 1 of 41 (2.4%) patients had isolated tumor cells, 21 of 78 (26.9%) patients had micrometastases, and 133 of 162 (82.1%) patients had macrometastases. The pooled sensitivity of intraoperative SLN frozen section analysis was 65% (95% CI, 51-77%) for macrometastases, micrometastases, and isolated tumor cells. When we excluded patients with isolated tumor cells, the pooled sensitivity increased to 72% (95% CI, 60-82%). CONCLUSION: SLN frozen section detects 65% of lymph node metastases compared with SLN ultrastaging and may prevent unnecessary radical surgery in some patients with early-stage cervical cancer.

Diagnostic accuracy of frozen section and patterns of nodal spread in high grade endometrial cancer: A secondary outcome of the SENTOR prospective cohort study.

OBJECTIVES: The study aimed to define the accuracy of intraoperative frozen section (FS) for the detection of metastases in sentinel lymph node biopsy (SLNB) and describe the pattern of lymph node (LN) spread and relation to molecular classifiers in patients with high-grade endometrial cancer (EC). METHODS: We performed a secondary outcome of clinicopathologic data from the Sentinel Lymph Node Biopsy versus Lymphadenectomy for Intermediate- and High-Grade Endometrial Cancer Staging (SENTOR) prospective cohort study evaluating SLNB in patients with clinical stage I high-grade EC (ClinicalTrials.gov ID: NCT01886066). The primary outcome was the sensitivity of FS of the sentinel lymph node (SLN) specimen, compared to a standardized ultrastaging protocol. Secondary outcomes included the pattern and characteristics of LN spread. RESULTS: There were 126 patients with high-grade EC with a median age of 66 years (range:44-86) and a median Body Mass Index (BMI) of 26.9 kg/m2 (range:17.6-49.3). FS was performed on surgical specimens from 212 hemipelves; SLNs were identified in 202 specimens (95.7%) and fatty tissue alone was identified in 10 specimens (4.7%). Of the 202 hemipelves in which SLNs were identified, 24 were positive for metastatic disease on final pathology. Initial FS correctly identified only 12, yielding a sensitivity of 50% (12/24, 95% CI 29.6-70.4) and a negative predictive value of 94% (178/190, 95% CI 89-96.5). A total of 24 patients (19%) had LN metastases: 16 (13%) had isolated pelvic metastases, 7 (6%) had both pelvic and para-aortic metastases and 1 (0.8%) had an isolated para-aortic metastasis. CONCLUSIONS: Intraoperative FS of SLNs in high-grade EC patients has poor sensitivity. Since isolated para-aortic metastases are rare, para-aortic lymphadenectomy may be omitted in patients in which SLNs were successfully mapped to the pelvis.

Confocal laser microscopy for assessment of surgical margins during radical prostatectomy.

OBJECTIVE: To evaluate the feasibility of confocal laser microscopy (CLM) for intraoperative margin assessment as faster alternative to neurovascular structure-adjacent frozen-section examination (NeuroSAFE) during robot-assisted radical prostatectomy (RARP). PATIENTS AND METHODS: Surgical margins were assessed during 50 RARP procedures in patients scheduled for NeuroSAFE. Posterolateral sections were cut and imaged with CLM and further processed to conform with the NeuroSAFE protocol. Secondary resection (SR) was performed in case a positive surgical margin (PSM) was observed with NeuroSAFE. Afterwards, the CLM images were non-blinded assessed for the presence of PSMs. The accuracy of both NeuroSAFE and CLM was compared with conventional histopathology. Agreement for detection of PSMs between NeuroSAFE and CLM was evaluated with Cohen's kappa coefficient. Procedure times were compared with a Wilcoxon signed-ranks test. RESULTS: In total, 96 posterolateral sections of RP specimens were evaluated for the presence of PSMs. CLM identified 15 (16%) PSMs and NeuroSAFE identified 14 (15%) PSMs. CLM had a calculated sensitivity, specificity, positive predictive value and negative predictive value of 86%, 96%, 80% and 98% respectively for the detection of PSMs compared to definite pathology. After SR, residual tumour was found in six of 13 cases (46%), which were all identified by both techniques. There was a substantial level of agreement between CLM and NeuroSAFE (κ = 0.80). The median procedure time for CLM was significantly shorter compared to NeuroSAFE (8 vs 50 min, P < 0.001). The main limitation of this study was the non-blinded assessment of the CLM images. CONCLUSIONS: Compared to NeuroSAFE, CLM is a promising technique for intraoperative margin assessment and is able to reduce the time of intraoperative margin assessment.