Liquid‐based cytology and cell block immunocytochemistry in veterinary medicine: comparison with standard cytology for the evaluation of canine lymphoid samples
Liquid-based Cytology (LBC) consists of immediate wet cell fixation with automated slide preparation. We applied LBC, cell block (CB) and immunocytochemistry to diagnose canine lymphoma and compare results with conventional cytology. Samples from enlarged lymph nodes of 18 dogs were collected and fixed in preservative solution for automated slide preparation (LBC), CB inclusion and immunophenotyping.
Two CB techniques were tested: fixed sediment method (FSM) and agar method (AM). Anti-CD79a, anti-Pax5, anti-CD3 and anti-Ki67 were used in immunocytochemistry. LBC smears showed better nuclear and nucleolar definition, without cell superposition, but presented smaller cell size and worse cytoplasmic definition. FSM showed consistent cellular groups and were employed for immunocytochemistry, while AM CBs presented sparse groups of lymphocytes, with compromised analysis. Anti-Pax-5 allowed B-cell identification, both in reactive and neoplastic lymph nodes.
Our preliminary report suggests that LBC and FSM together may be promising tools to improve lymphoma diagnosis through fine-needle aspiration.
Introduction
Cancer is the main cause of death in North American adult dogs and lymphomas are among the most common types of dog tumors, affecting animals of all breeds and ages.
Fine-needle aspiration biopsy (FNAB) is a fast and cost-effective tool for the initial diagnostic planning of neoplastic diseases in general4 and plays a pivotal role in lymphoma diagnosis, as well in monitoring disease remission and relapse.
However, although some studies have described high specificity and sensitivity of FNAB for differentiating lymphoma from lymphoid hyperplasia in adequate samples, an inadequate technical procedure may yield a low-quality smear, with artefacts such as thick smears, contamination, improper/delayed fixation, low cellularity and cell disruption, thus preventing adequate morphological analysis. Fine-needle aspiration may be performed by a clinician, avoiding anesthesia and additional costs to the owner, while histopathological analysis requires a more invasive procedure. Many techniques are being applied in order to enhance lymphoma diagnosis through fine-needle aspiration, such as flow cytometry, gene profiling, PCR for antigen receptor rearrangements (PARR) and immunocytochemistry.
Considering these issues, we propose to introduce liquid-based cytology (LBC) and cell block (CB) as adjunct techniques for the cytological diagnosis of lymphoma. LBC has been used in human medicine since the mid-1990s, and consists of immediate transfer of harvested cells (by exfoliation or aspiration) to a preservative fluid with a decreased number of inadequate samples per harvest. In addition, the samples are suitable for multiple slide preparation, as well as further tests such as biomolecular tests, CB and immunocytochemistry.
LBC has been used for several non-gynecological specimens in human medicine, such as cavity body fluids, urine, cerebrospinal fluid, sputum, bronchial washes and aspiration biopsies, including lymph nodes. However, to our knowledge there are no LBC studies in veterinary medicine, despite the paramount importance of cytopathological diagnosis in clinical practice.
CB technique consists of sedimenting fixed cells for paraffin wax block embedding. This technique can be used to preserve the residual sample from LBC in the same manner as it is already performed in human medicine. Furthermore, CB increases the sensitivity in malignancy detection and reduces false-positive results.
In this study, we compared LBC samples with traditional air-dried cytology samples from dogs with lymphoma or reactive lymphoid hyperplasia, regarding morphological features, and compared two methods of CB technique: fixed sediment method (FSM) and agar method (AM). We also performed immunocytochemistry technique in CB samples for lymphoma immunophenotyping and determination of proliferation index.
Authors: N. C.C. A. Fernandes, J. M. Guerra, R. A. Réssio, D. G. Wasques, D. Etlinger-Colonelli, S. Lorente, E. Nogueira, M. L. Z. Dagli
Source: https://onlinelibrary.wiley.com/
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