Liquid‐based cytology and cell block immunocytochemistry in veterinary medicine: comparison with standard cytology for the evaluation of canine lymphoid samples

Abstract

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.

Cytodiagnostics of canine lymphomas – possibilities and limitations

“…Ki67 ICC has also proved to be an excellent method of differentiation between non-neoplastic and neoplastic proliferation of lymphatic tissue (Bauer et al 2007, Fernandes et al 2015. Moreover, Fournel--Fleury et al (1997b) have confirmed that immunocytochemistry using Ki67 antibodies is a useful method of determination of proliferative activity of lymphoma cells, and have revealed high concordance between the results of Ki67 immunocytochemistry and immunohistochemistry performed on histological samples of lymphoma tissues.…”

“…In canine oncology it is especially valuable for diagnosing the most common subtypes of lymphomas (Fournel-Fleury et al 1997a, Fournel-Fleury et al 2002, Comazzi et al 2014. Recently, many authors have recommended using cytology as a sufficient diagnostic method for canine lymphomas, especially if supported by immunocytochemistry and/or flow cytometry (Messic 2008, Regan et al 2012, Martini et al 2013, Comazzi et al 2014, Curran and Thamm 2015, Fernandes et al 2015, Jankowska et al 2015. The great advantage of this method is the possibility of performing immunocytochemical staining.…”

“…Since the immunophenotype of neoplastic cells influences both treatment protocol and prognosis, immunophenotyping is recommended as an obligatory diagnostic method during characterization of canine patients with lymphoma by veterinary oncologists in the USA (Regan et al 2012). ICC using other antibodies (CD4, CD8, CD21, CD45 and others) allows for precise subtyping of lymphoma cells and thus confirmation of less common or unusual types of tumors (Dzimira 2007, Martini et al 2013, Pawlak et al 2014, Fernandes et al 2015.…”

“…The updated Kiel classification (uKc) adopted for dogs is the second classification system commonly used for canine lymphoma subtyping (Fournel-Fleury et al 1997a, Fournel-Fleury et al 2002, Sozmen et al 2005, Ponce et al 2010, Jankowska et al 2015, Fernandes et al 2015. This system is based on neoplastic cell morphology and immunophenotype, with regard to similarity to normal lymphocytes in various stages of maturity.…”

Pax5 as a potential candidate marker for canine B-cell lymphoma

“…Pax5 staining was used for B-cell identification in a complicated case report on multicentric B-cell lymphoma with neurolymphocytosis (Schaffer et al 2012). Pax5 was also employed in a study of inflammatory processes: spirocercosis-induced nodular formation in dogs (Dvir et al 2011), and was also used in an immunocytochemistry by liquid-based cytology and tissue transfer technique for immunophenotyping in canine non-Hodgkin lymphoma (Stone and Gan 2014;Fernandes et al 2015). The sensitivity and specificity of Pax5 immunohistochemistry for B-cell lymphomas when compared to heteroduplex polymerase chain reaction for antigen receptor rearrangements of IgH primer sets for B-cells, were 48% and 100%, respectively (Sirivisoot et al 2016).…”