| Abstract|| |
Context: Liquid-based cytology. Aims: Utility of liquid-based cytology (LBC) was compared to conventional smear cytology in ultrasound-guided fine-needle aspirates of abdominal masses. Settings and Design: This was a prospective comparative study conducted in collaboration with surgery and pediatrics surgery departments of our institute. Subjects and Methods: Thirty patients presenting with evidence of abdominal mass were enrolled for the study and underwent fine-needle aspiration cytology. The material was processed for the preparation of conventional smears and residual material was rinsed into cytolyt for LBC by Thin Prep method and into cell block fluid. The smears prepared from both the methods were compared by two independent and experienced pathologists for adequacy, cellularity, architectural pattern, cytoplasmic preservation, nuclear preservation, and background. Results: Cellularity was frequently higher in the conventional smears than on Thin Prep slides (P value = 0.025). Recognition of architecture was better on the conventional smears (P value = 0.001). Cytoplasm was better preserved on the conventional smears (P value = 0.001) but difference in the preservation of nuclear details was not statistically significant on slides prepared from both the techniques. The background of Thin Prep slides is significantly cleaner than direct smears (P value = 0.001). Non epithelial elements such as mucin and neurofibrillary tangles were better preserved on direct smears (P value = 0.001) but diagnostic accuracy of both the methodologies showed no statistically significant difference (P value = 0.226). Conclusions: The Thin Prep technique utilizes expensive equipment and reagents. It also generates certain morphological artefacts in slides with which a cytologist needs to get familiar. When used in isolation, it may not consistently provide any added advantage in the diagnosis of such lesions and should be used as an adjunct to conventional smears. It may be preferred in situations where material has to be transported for processing or is required for ancillary tests.
Keywords: Abdominal masses, FNA, liquid-based cytology, Thin Prep
|How to cite this article:|
Choudhury M, Pahwa AR. Comparison of liquid-based cytology with conventional cytology in the evaluation of abdominal masses. Indian J Pathol Microbiol 2020;63:73-7
|How to cite this URL:|
Choudhury M, Pahwa AR. Comparison of liquid-based cytology with conventional cytology in the evaluation of abdominal masses. Indian J Pathol Microbiol [serial online] 2020 [cited 2021 Jun 14];63:73-7. Available from: https://www.ijpmonline.org/text.asp?2020/63/1/73/277377
| Introduction|| |
Liquid-based preparations were originally developed for application to Cervical smears but have progressively gained favor when applied to both nongynecological cytology samples and fine-needle aspirate cytology.
Liquid-based preparations have several preparatory, screening, and diagnostic advantages including uniform collection procedures, avoiding hazards of needle handling required during conventional preparation, and easy transportation to the laboratory, making them an appropriate alternative to the conventional cytology. It requires immediate liquid fixation, which preserves morphology and doesn't lead to any air-drying artefact. The residual material can be used to process multiple slides, cell block for ancillary tests such as immunocytochemistry, and other molecular tests, namely, PCR, flow cytometry, and DNA ploidy analysis.
In the recent past, studies have compared the traditional smear technique to liquid-based, thin-layer technology in the evaluation of gynaecologic material. These studies determined that liquid-based methods increase the detection of intraepithelial lesions and infectious agents and decrease the proportion of unsatisfactory diagnoses. Studies have also been performed on nongynaecologic material with variable results.
However, in order to justify the use of such a technique, which adds substantial cost to existing procedures, a thorough unbiased investigation needs to be undertaken to determine its diagnostic value. Several studies have conflicting results regarding the quality and adequacy of specimen preparation of FNA smears by this technique.,
The two FDA approved liquid-based cytology (LBC) techniques for both gynaecological and nongynaecological cytology samples are Thin Prep (CYTYC Co., Marlborough, MA) and Tri Path PREP (TriPath, Inc., Burlington, NC). There are many manual techniques also such as SYNERMED GLUCYTE®, PAPSPIN (Thermoshandon), CYTOSCREEN (Seroa, Monaco), TURBITEC (Labonord, Templemars, France), and others.
The direct smears are prepared by manual smearing of the sample on a slide because of which there is lack of consistency and little control over critical sample components such as cellular morphology, clarity, density, and uniformity. Cellular distribution on slides tends to be uneven and cells are likely to get damaged by air drying and get obscured by blood, mucus, and overlapping cells.
There are many studies in the literature assessing utility of LBC techniques in the fine-needle aspiration cytology (FNAC) of breast and thyroid but there are relatively few studies on their role in abdominal aspirates.,
This study was undertaken to evaluate the role of LBC and compare it with that of conventional smear technique in the aspirates of abdominal masses.
| Subjects and Methods|| |
The comparative study was conducted over a span of 15 months with the approval of Institutional ethical committee. Thirty patients including those from pediatric ward presenting with clinically palpable and/or radiologically detectable abdominal lesions were included. Ultrasound-guided fine-needle aspirate and cell block were collected. In some cases, the aspirate was obtained postoperatively from the surgical specimen before immersing in formalin such as kidney or bladder tumors.
Exclusion criteria were the masses with inflammatory, traumatic etiology, and the cases where diagnosis could not be confirmed due to the lack of histopathological correlation.
The aspirates were spread on clean glass slides and were air-dried as well as fixed in 95% ethanol for staining.
The same syringe was rinsed into liquid-based collection vial, which contains proprietary methanol-based preservative solution. The material was processed in Thin Prep 2000®, one smear was stained with Papanicolaou stain and one with hematoxylin and eosin stain.
Processing of samples for LBC using the Thin prep2000 was carried out as per manufacturer's instructions. First, the cells collected in Cytolyt were centrifuged at 1000 rpm and then transferred into Preservcyt. After 15 min, the collection vial was then inserted in the Thin Prep 2000 processor and slides were prepared using membrane transfer technique on Programme 2.
Smears were stained by standard Papanicolaou, Hematoxylin eosin, and Giemsa stain. After preparation of slides from both the methods, they were compared for cellularity, cellular architecture, maintenance of nuclear detail, cytoplasmic integrity, background including presence of matrix/stromal material and diagnostic accuracy. Cell block or biopsy correlation whichever available was done. The slides prepared with each of the techniques were examined by two independent and experienced pathologists.
The aim of this study was to compare both the techniques based on the assessment of the parameters mentioned.
| Results|| |
Out of 30 cases, six belonged to the pediatric age group and were less than 15 years of age. Twenty-four cases were adults with age ranging from 40–70 years; a mean age is 56 years.
Both the smears were examined by two different observers and graded using a semiquantitative scoring system as in [Table 1]. The final diagnoses of all the 30 cases are enlisted in [Table 2]. Based on each parameter, the cases were categorized into three groups as in [Table 3].
|Table 1: Semiquantitative scoring of cytological features on conventional and liquid-based preparations|
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|Table 3: Original: categorization and comparison of CS and TP after grading parameters on examining smears|
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All but three cases were diagnosed on the basis of tissue biopsy. Three cases were instead diagnosed on cell block preparation—Neuroblastoma, Yolk sac tumor, and Hepatoblastoma.
Using the histologic/cell block diagnosis as the gold standard, the number of cases correctly diagnosed on Conventional and the Thin Prep slides were 29 (96.67%) and 25 (83.33%), respectively. The difference in the diagnostic accuracy between the two methods was not statistically significant (P value = 0.226).
The result of comparison of both the methods with regard to cytologic features was as follows:
Cellularity, Architecture, Cytoplasmic Integrity—conventional smears superior to Thin Prep (P = 0.025, <0.001, <0.001).
Adequacy and Nuclear details—Difference not statistically significant (P = 0.112, 0.091).
Background material–Thin Prep superior to conventional smears (P < 0.001).
| Discussion|| |
The smear technique has been universally used for the processing of fine-needle aspiration material. After slide preparation for routine staining, the material in the syringe and needle is either discarded along with the syringe or needle, or is rinsed in various solutions, most commonly in RPMI-1640 medium or recently in Preservcyt™. The material in the rinse solution is then processed either as cell block, cytospin, or for monolayer preparation.
Thin layer preparations especially those prepared by automated techniques may lead to some changes in the cellular and background morphology considering which cytopathologists need to be trained in diagnostic practices in thin layer cytology.
The present study was undertaken to compare the liquid-based technique with conventional smears in the evaluation of abdominal masses.
The cellularity on conventional and Thin Prep slides was equivalent in 53.3% (18), better on conventional in 40% (10), and better on Thin Prep in 6.6% (2) cases. These findings correlated with those of Warren et al. wherein cellularity was equivalent on both preparations in 54%, better on conventional in 34%, and better on Thin Prep in 12% cases.
Conventional smears were adequate for evaluation in 100% cases. Smears from aspirates of 26 patients (86.7%) prepared by Thin Prep were adequate to make a diagnoses. This maybe due to the fact that cells got diluted in collection medium in Thin Prep smears. Nasuti et al. observed that a diagnosis could be made on 63% cases based on Thin Prep slides while 21% were less than optimal and16% were unsatisfactory for evaluation.
Salhadar et al. observed that 98% of conventional smears and 64% of Thin Prep slides were satisfactory and adequate for evaluation from material obtained from 50 FNA specimens.
Architectural pattern of the different lesions was equally well recognized on the conventional smears and Thin Prep in 40% (12) and better recognized on conventional smears in 60% (18) cases [Figure 1]. Thin Prep was not better than conventional smears in any case with regard to architecture. These findings were consistent with those of Leung et al. who observed that recognition of architecture was equivalent on both conventional and Thin Prep slides in 24%, better on conventional in 58%, and better on Thin Prep in 18% cases.
|Figure 1: FNAC Conventional Pap stain—Smear from Papillary RCC showing well formed papillae (100×)|
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The background was significantly cleaner on Thin Prep slides in 76.6% (23) cases and was equivalent on both the preparations in 23.3% (7) cases. None of the conventional slides showed a cleaner background than Thin Prep slides. The difference was statistically significant. These findings were in agreement with findings of Leung et al. who noted that background was better on ThinPrep in 75%, and equivalent on both ThinPrep and conventional in 21%, and better on conventional smears in 4% cases.
These findings were similar to those of Sung Hoon et al. who investigated the usefulness of LBC on EUS-FNA. They found that although the background was cleaner on LBC, the cellularity was decreased.
However, it is important to note that in certain cases, background material gave clues to the diagnosis. The presence of fibrillary material suggestive of neuropil on conventional smears was helpful in diagnosing one case of neuroblastoma [Figure 2] and the presence of mucin was suggestive of the type of colonic and gastric adenocarcinoma in two cases. These tend to decrease or get lost in processing in liquid-based preparations as shown in [Figure 3].
|Figure 2: FNAC Conventional Pap stain—Neuroblastoma showing identifiable rossette with central neuropil (400×)|
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|Figure 3: FNAC LBC—Neuroblastoma Pap stain cluster showing frayed edges, lysed blood in background, loss of neuropil (400×)|
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The utility of LBC technique varied in FNA from different sites according to Tripathy et al. who observed that in case of salivary gland lesions, the mixed salivary tumor was better visualized in the conventional smear because of the preservation of the stromal component. Cystic neoplasms of the salivary glands were better appreciated in LBC smears because of concentration. They also reported the loss of stromal material on FNA from fibroadenoma breast.
The cytoplasm was better preserved in conventional preparations in 73.3% (22) cases and equally well preserved on both in 26.6% (8) cases. None of the Thin Prep slides showed better cytoplasmic preservation than conventional smears.
The nuclear details were equally well preserved on both conventional smears and Thin Prep in 63.3% (19), better on conventional in 33.3% (10), and better on Thin Prep in 3.3%(1) cases. This difference was not statistically significant.
Hence, from our study, we observe that overall cytomorphological preservation was better on conventional slides. This refutes the findings of Lee et al. who stated that cytologic preservation was better on Thin Prep in 76%, better on conventional in 3%, and equivalent on both in 21% cases.
Michael et al. noted that except bloody background, conventional smears were equal in quality in 54%, better in 34%, and inferior to Thin Prep in 11.6% cases.
Diagnostic accuracy of conventional slides was calculated as 96.7% compared to 83.33% with the Thin Prep technique. Nasuti et al. noted a reduction in the number of cases definitively diagnosed as malignant (54% reduced to 33%) on reviewing Thin Prep slides alone.
Our findings were similar to a study on diagnostic value of Liquid-Based Cytology With Fine Needle Aspiration Specimens for Cervical lymphadenopathy by Bandoh et al. (2016) where the diagnostic rate of pathological malignancy using Conventional cytology was 52.4% and that of LBC was 50.4%.
Lee et al. compared the results of LBC and the conventional smear method when performing endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) for lesions of suspected pancreatic malignancy with a prospective study on 58 patients.
They concluded that liquid-based preparation was less accurate than conventional smears when performing pancreatic EUS-FNA but it might serve as a good complement to conventional smears if blood contamination is profound.
We observed that morphologically the cells in all LBC smears appeared relatively smaller and were arranged in tighter three-dimensional clusters, with condensed cytoplasm and fraying of edges in some slides. Cells on LBC preparations also tend to exhibit more hyperchromatic nuclei than seen on conventional smears as shown in [Figure 4].
|Figure 4: FNAC LBC Pap stain—Papillary RCC showing a papilla with cells in tight overlapping cluster (100×)|
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These observations were in agreement with those of Garbar et al. who also reported that cells on LBC of FNA lymph nodes show retracted cytoplasm and hyperchromatic nuclei.
A cytopathologist also needs to get familiar with slides prepared by this technique due to the artefacts induced by the processing technique. It might be more effective in laboratories where the pathologist does not have any control over slide preparation of aspirates like remote areas where a clinician needs to collect the aspirate and in cases where ancillary tests are required for evaluation.
In conclusion, it is an expensive technique for routine use and has less diagnostic utility when used alone than in conjunction with conventional techniques.
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Conflicts of interest
There are no conflicts of interest.
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Archna R Pahwa
A002, Raheja Atlantis, Sec 31, Gurugram - 122 001, Haryana
Source of Support: None, Conflict of Interest: None
[Figure 1], [Figure 2], [Figure 3], [Figure 4]
[Table 1], [Table 2], [Table 3]