Liquid Based Cytology

LBC have been developed to improve the detection of cervical cancer and its precursors and reduce the rate of false negative results from conventional cervical smear tests. In several countries liquid based cytology is replacing conventional smear tests. A thorough economical evaluation is needed for this new technique (LBC) to be implemented in our country as a mass screening and also human papilloma virus testing as complementarily to conventional smear testing should be further evaluated in clinical research.

Screening for cervical cancer has resulted in the reduction of incidence of cancer cervix and its complications. Pap smear has been used for the screening purpose since 1940’s but studies have shown that the sensitivity of a single Pap test is low and it has high false negative rate .This is said to be due to improper sampling and ineffective interpretation. In Pap smear only 20% of the scraped cells in the Ayer’s spatula are transferred to the slide while smearing, which is done manually. Therefore alternative screening methods which overcome these pitfalls in Pap smear is developed, one such method is Liquid Based Cytology where all the scraped cells in the cyto brush are transferred in the liquid transport medium to the laboratory where it is processed and a uniform thin layer of smear is prepared in the laboratory than manually. This study is done for evaluating the performance of Liquid Based Cytology and for doing comparative study of the sensitivity of LBC and the conventional Pap method.

Differential cell counts (DCC) of induced sputum samples have been shown to be useful in the management of patients with moderate–to-severe asthma(1) and in the diagnosis of eosinophilic bronchitis(2). Unfortunately, sputum processing is time consuming(3) taking approximately 4 hours and a meta-analysis concluded that its routine use could not be justified due to the technical expertise required(4). We aimed to establish if a simplified method using routine liquid based cytology (LBC), used in histopathology laboratories throughout the NHS, could provide similar results.

Liquid based cytology has evolved as a newer method in the field of fine needle aspiration cytology. The introduction of liquid based technique tries to overcome the limitations faced by conventional method. Liquid based smears are easier to screen and less time consuming due to spread of cells in monolayer with a clean background. The cellular morphology is always well preserved. Only a small amount of is used for LBC preparation and the remaining solution is stored in appropriate preservative solution for few months at an appropriate temperature for future studies.

important public health problem. The cellular changes in cervix and intraepithelial lesions can be detected many years before the patients present with frank invasive carcinoma. So, cervical screening programs were introduced worldwide. For many years , Conventional PAP smears were used for screening. Though it led to drastic reduction in number of cervical carcinoma cases, it had high false negativity. So, newer methods like Liquid based cytology were introduced. This study was undertaken to compare Liquid based cytology with Conventional PAP smear and to correlate the results with biopsy obtained from the same patient. This study was done on randomly selected 100 patients attending the Pilot screening project at Department of Obstetrics and Gynaecology , Thanjavur medical college ,

both as an adjunct to cytological screening and in pri- mary screening for cervical dysplasia. Many efforts have been made to develop methods, which would enhance the sensitivity and specificity of the Papanicolaou smear. From various researches the evolution of liquid-based gynecologic specimen was resulted. The researchers ar- gue that liquid-based preparations outperform conven- tional method smears because of improved fixation; standardizations of cell transfer and decreased obscuring factors. Another advantage of liquid based cytology is that the transport media containing the remaining cells could be stored and used for HPV DNA testing. The re- searchers also point out that, in direct smears, the cells may not transfer in a representative way and that up to 90% of the scraped materials from the cervix may be discarded. As with liquid-based collection, the sampling will be operator dependent and variation is very less likely, since the laboratory controls the processing. And with proper sampling liquid based cytology has the ad- vantage of being suitable for high-risk HPV testing and could reduce unsatisfactory specimens from 4.1% to 2.6% [8].

Cytology for the detection of endometrial malignancy has re-emerged, since more elaborate methods for sam- pling and storage of cells have been introduced. As a result, in many studies cytology has fewer inadequate samples than biopsies, while it remains less intervene- tional and more tolerable [19,29,30]. In the limited num- ber of the samples we examined a single sample which was inadequate, while all women found the sampling method very tolerable. Introduction of liquid based cy- tology has limited the factors reported to hamper safe cytology diagnosis [17], while many report extremely high diagnostic efficacy of liquid based cytology using current sampling and preparation methods [21,22]. In our samples, the only missed carcinomas were one that was inadequately sampled, a serous carcinoma and a single carcinosarcoma that are extremely rarely found. As a result high sensitivity, specificity and PPV were identi- fied (Ta- ble 3). Even though the same diagnostic criteria of conventional cytology can be applied to liquid based cytology [20], hyperplasia with atypia was found to be the most controversial category among observers [31], while it has been propose that different commercial liq- uid based preparation methods may result in morpho- logical alterations [32].

Forty-five BD SurePath-preserved specimens and 45 Hologic PreservCyt-preserved specimens were selected from a database of previously tested samples with the BD hrHPV-GT assay. The sam- ples were originally collected from cytology labs as deidentified residual vial specimens after the liquid-based cytology slides had been prepared. Thus, the samples were originally exposed to am- bient storage temperatures (15 to 30°C) in accordance with the respective manufacturers’ recommendations during collection, transit to the cytology laboratory, and final transfer of the residual material to long-term storage at 2 to 8°C. Upon receipt, approxi- mately 16% of the SurePath specimens and 31% of the PreservCyt specimens were beyond their recommended storage dating, and at the time of first testing, all samples had expired. The original test- ing was performed in the fall of 2009, and each specimen tested was positive for at least one HPV type. The second test was per- formed in the spring of 2012, at which time the mean age of both SurePath and PreservCyt specimens was 2.6 years. All 90 samples were collected from different individual patients but were pur- posely selected so as to provide a balanced representation of high- risk types detected by the assay for both medium types (Table 1). One SurePath sample was removed from the study due to a label- ing error. A subset of samples from the same collection was also analyzed by endpoint PCR for detection of the human GAPDH (glyceraldehyde-3-phosphate dehydrogenase) gene.

The Hybrid capture II HPV DNA test (Digene/Abbott, Maidenhead) was performed on liquid based cytology specimens. The RNA probe mix for the detection of high risk HPV types 16,18,31,33,35,39,45,51,52,56, 58,59 and 68 was used according to the manufacturer's instructions. Signal amplification is based on immunocapture of DNA/ RNA hybrids and subsequent EIA with a chemilumines- cent reporter system. The results are given as relative light unit (RLU) ratio. As the RLU ratio is the ratio of light emit- ted by the specimen to the light emitted from the mean RLU of triplicate positive control specimens containing 1 pg/ml of HPV DNA (5000 copies of HPV genome)it is a semiquantitativ estimate of viral load in the specimen. Cost comparison

Abstract: Histiocytic necrotizing lymphadenitis (HNL; Kikuchi-Fujimoto disease) is a rare benign disorder. The di- agnosis of HNL is established on recognizing the characteristic histologic findings from biopsy of the enlarged lymph nodes. Though diagnosis of HNL by fine-needle aspiration (FNA) was reported, the characteristic fine-needle aspiration cytologic features with conventional cytology and a liquid based cytology test (LCT) have not been well documented. In this study, 42 cases of suspicious necrotic lymph nodes were subjected to cytology and biopsy diag- nosis. The lymph nodes were aspirated using a 10 mL disposable syringe with the percutaneous ultrasound guided. Samples were used for conventional cytology and LCT. Among 42 cases of suspicious necrotic lymph nodes, 37 of cases were histologically confirmed as HNL; 3 of cases were hyperplasia of lymphoid tissue; 1 case was tuberculosis of lymph node, and 1 case was classical Hodgkin lymphoma (nodular sclerosis type). 31 out of 37 (83.8%) cases of HNL were diagnosed by conventional cytology, 33 out of 37 (89.2%) were diagnosed by LCT. Our results indicate that no significant difference on accuracy rate between conventional cytology and LCT, but LCT has its advantages in the diagnosis of HNL.

LBCs are a suitable biospecimen for high-throughput contexts. However, cell numbers in individual samples can vary widely and the sample can contain HPV-infected and uninfected cells. Moreover, the cells in the sample can represent a range of disease stages if these are present across an individual cervix. Consequently, percentage positivity for either biomarker assessed in the present study likely gives only an estimate of their levels in-situ. Our choice of cut-o ff values for MCM and E4 could be inappropriate given the small number of samples tested. It will be useful in a future study with greater numbers of samples to compare data with and without cut-o ff values. Despite these caveats, as predicted by the previous tissue-based study, high MCM detection was associated with CIN2+ and E4 association with < =CIN1 was close to signi fi cance. E4 expression levels can be very variable in CIN1 lesions [12] and, as a cytoplasmic protein it may be more likely than a nuclear protein to leach out of LBC cells during storage and processing. How- ever, the low numbers of samples in the study will impact the sig- ni fi cance of the E4 results. Some E4 was detected in histopathologi- cally-defined “normal” samples. These were HR-HPV positive and therefore likely represent transient infections. A further study using a larger cohort is required to validate E4 as a biomarker of clinically insignificant disease in LBC samples, or conversely, that low or no E4 expression is a biomarker of CIN2 + . A previous study found that de- tection of p16 but no detection of viral L1 protein was su ffi cient to predict CIN3 [25]. It would be useful to investigate a quadruple bio- marker matrix incorporating p16, MCM, E4 and L1 in an LBC-based cervical disease risk-strati fi cation test. However, biomarker analysis is not straightforward. For example, although p16 and MCM are bio- markers of clinically significant disease, the former is found in CIN1 and CIN2+, albeit with a different distribution, and is present in senescent cells while the latter can be present during metaplasia or in fl ammation. Detection of the viral biomarkers E4 and L1 seems to more restricted to ≤ CIN1 [12,13]. Thus combinations of markers, especially when particular patterns are pathognomonic of disease, should o ff er clear benefits.

a liquid fixative containing alcohols and/or formalde- hyde and subjected to cytological tests (liquid-based cytology; LBC). In general, different types of thyroid tumors can be easily differentiated by histopathological diagnosis. However, there are exceptions; for example, distinguishing between metastatic and non-metastatic minimally invasive (MI)-follicular thyroid carcinomas (FTCs) is currently difficult by any pathological modali- ties. Thus, it is necessary to establish a method for the molecular diagnosis of thyroid tumor to substitute or complement the histopathological method.

has fallen from 13.6% to 1.9% [6]. Thus, LBC has im- proved unsatisfactory conventional Pap smear rates and give significant benefits to colposcopic examinations. Moreover, LBC technique shortens laboratory turn- around times. The superiority of the quality of LBC in comparison with those of conventional Pap smears has been described [2,7,8]. The sensitivities of the conven- tional Pap smear and LBC tests are estimated to be 70% - 80% and 85% - 95%, respectively [9]. LBC is currently recommended for cervical cancer screening [10] with a major advantage of allowing ancillary techniques such as those used in immunocytochemistry and molecular bio- logy [11-17]. Cell blocks made from cell or tissue rem- nants of LBC can also be used for immunocytochemistry or immunohistochemistry of specific biomarkers to ac- curate diagnosis of malignancy [16-19]. In addition, LBC has currently been used for non-gynecologic cytology [15,17,19-22].

I solemly declare that this dissertation titled “Comparison between liquid based cytology and conventional cytopreparatory methods in body cavity fluids” is the original and bonafide work done by me under the guidance of Dr.S.Mary Lilly, M.D., Head of Department and Professor,Department of Pathology at the Government Stanley Medical College & Hospital, Chennai -600 0001, during the tenure of my course in M.D.Patholgy from May-2012 to April 2015 held under the regulation of the Tamilnadu Dr.M.G.R. Medical University, Guindy,Chennai-600032

Objectives: (1) To evaluate the various causes of unhealthy cervix in Kashmiri women. (2) To assess the utility of colposcopy and cytology in detecting the various causes of unhealthy cervix. (3) To correlate the findings of cytology and colposcopy with each otherand with histopathology. Methodology: 200 women attending gynecology OPD with clinicallyunhealthy cervix were subjected to evaluation by liquid- based cytology, colposcopy and directed biopsyand the findings were noted. Results: Majority of women were in the age group of 30-39 years. Commonest complaint was vaginaldischarge in 50% cases followed by menorrhagia in 19.5% women18% had abnormal discharge in 50% cases followed by menorrhagia in 19.5% women, 18% had abnormal LBC findings with 11% women had LSIL, 5.5% had HSIL, and 1.5% had squamous cellcarcinoma (SCC). 32.5% had abnormal women had LSIL, 5.5% had HSIL, and 1.5% had squamous cell carcinoma (SCC). 32.5% had abnormal colposcopic findings, 21% had low grade lesion and 11.5% had high grade lesion. In our study, on correlating LBC findings with histopathology, out of 8.5% women with ASCUS on LBC, 4% had positive histopathological findings. Out of 5% patients with LSIL on LBC, 3.5% were having positive histopathologicalfindings. Out of 3% patients with HSIL on LBC, 1.5% were positive on histopathological examination. On correlating colposcopic findings with histopathology, 5.5% patients which appeared normal on colposcopic evaluation had dysplasia on histopathology, 15.5% women with low grade lesion on colposcopy had

Abstract: Background: Controversy exists about the diagnostic value of liquid-based cytology (LBC) compared to conventional smears (CS). Most prior studies of LBC were performed using ThinPrep system. Few studies have ever compared the adequacy rate of SurePath with conventional smears. Methods: We performed a prospective com- parison of LBC using SurePath with CS in 304 thyroid nodules. Four needle sticks constituted a single nodule FNA, with 2 passes used for CS while the other 2 passes were used for SurePath. Cytopathologists separately read all samples, and all slides were reported using the Bethesda system for reporting thyroid cytology. The adequacy rate was compared between the CS and SurePath groups. Results: The adequacy rate for all solid nodules was 78.2% in CS group, significantly higher than 68.0% in the SurePath group (P=0.006). No significant difference was seen for mixed or cystic nodules. The adequacy rate using a combination of CS and SurePath in solid nodules was 86.4%, significantly higher than 78.2% in CS group (P<0.001). When excluding nodules less than 1 cm, the adequacy rate of CS for solid nodules was 83.5%, significantly higher than 71.3% in SurePath group (P=0.02). The adequacy rate of combination of CS and SurePath was 91.3% for solid nodules, significantly higher than 83.5% in CS group (P=0.04). Conclusion: Our study showed that LBC using SurePath is not superior to conventional smears. However, a combina- tion of both SurePath and CS may yield the most favorable adequacy rate compared to either process separately.

Thyroid nodule is a common problem with high clinical prevalence in women ranging from 5.3 to 6.4 % (Ratour et al, 2013). Benign nodules outnumber the malignant one, but the risk of malignancy is to be evaluated preoperatively for (Gita Jayaram, 2012) Fine needle aspiration cytology (FNAC) is a simple, rapid, minimally invasive, cost effective procedure which plays an important role in evaluation of patients with thyroid pathology. FNAC is the most o separate benign nodules from malignant nodules which triages the patients of malignant Syed Z. Ali and ). Thyroid cytology reporting needs the communication between pathologist, treating physician or surgeon and radiologist. Previously pathologists were using

Breast cancer is known to be a heterogeneous disease .So tumors with the same clinicalpathological characteristics tend to exihibit diversity in disease behavior, response to therapy and prognosis. Gene expression profiling studies is supposed to be the gold standard method to subtype tumors based on molecular features.. As gene expression profiling from formalin-fixed, paraffin-embedded samples is not readily available at present ,IHC surrogate panels of estrogen receptor (ER), progesterone receptor (PR), HER2 and ki-67 can be readily used to subtype the malignant tumors of breast. (Nielsen et al., 2004; Livasy et al., 2006; Carey et al 2007; Hugh et al., 2007; Cheang et al., 2008).

Sixty two patients with clinically and/or radiologically detected intra-abdominal masses diagnosed on FNAC in the Department of Pathology during a 21 month duration in a tertiary care institution form the material for this study. Consent for this study was taken from the ethical committee of the institute. The intra-abdominal site of the lesions sampled included the liver, the retroperitoneum, abdominal lymph nodes, intestine, pancreas, stomach, kidney, omentum and mesentery. Pelvic masses were excluded from the study. Clinical data with emphasis on history, physical examination and relevant investigations were analysed thoroughly and discussed with the referring physician. Patients with severe coagulopathy and cases like haemangioma and hydatid disease of liver were considered as contraindications for FNAC in view of possible serious complications. The procedure of FNA was explained to the patient seeking an informed consent and complete co-operation for the procedure. The mass under investigation was evaluated by USG and the path of FNA providing the safest and easiest access to the lesion was chosen. Under aseptic precautions, FNAC was performed using 20 or 22 gauge, 90 mm spinal needle attached to a 10 ml disposable syringe. The prepared smears were stained with Papanicolaou and Haematoxylin and Eosin stains (H&E) stains. Periodic acid-Schiff, Gram’s and Ziehl-Neelsen stains were done wherever required. For CB analysis, all macroscopically visible tissue fragments in the aspirated material were collected in 10% formaldehyde solution and sedimented by centrifugation. Keywords: Abdominal mass, Cytology, Cell block histology, Ultrasound

In reviewing the clinical records, we considered leptomen- ingeal metastases to be present if positive results of CSF cyto- logic examination were obtained either 14 days before or 14 days after MR imaging of the brain. We considered leptomen- ingeal metastases to be absent if there was no clinical evidence of leptomeningeal disease with follow-up of ⱖ2 months and no positive cytology. The clinical criteria called for no new or progressive neurologic deficits for ⱖ2 months after MR imag- ing. Negative CSF cytology is not adequate because even three separate lumbar punctures can have a false negative rate of ⱕ10% (11). Additionally, none of these patients was being treated for leptomeningeal metastatic disease.