Utilization of Cell-Transfer Technique for Molecular Testing on Hematoxylin-Eosin–Stained Sections: A Viable Option for Small Biopsies That Lack Tumor Tissues in Paraffin Block

Abstract

Context.-In some instances the standard method of doing molecular testing from formalin-fixed, paraffinembedded block is not possible because of limited tissue. Tumor cell-enriched cell-transfer technique has been proven useful for performing immunocytochemistry and molecular testing on cytologic smears.Objective.-To establish the cell-transfer technique as a viable option for isolating tumor cells from hematoxylineosin (H&E)-stained slides.Design.-Molecular testing was performed by using the cell-transfer technique on 97 archived H&E-stained slides from a variety of different tumors. Results were compared to the conventional method of molecular testing.Results.-Polymerase chain reaction-based molecular testing via the cell-transfer technique was successfully performed on 82 of 97 samples (85%). This included 39 of 47 cases for EGFR, 10 of 11 cases for BRAF, and 33 of 39 cases for KRAS mutations. Eighty-one of 82 cell-transfer technique samples (99%) showed agreement with previous standard method results, including 4 mutations and 35 wild-type alleles for EGFR, 4 mutations and 6 wild-type alleles for BRAF, and 11 mutations and 21 wild-type alleles for KRAS. There was only 1 discrepancy: a cell-transfer technique with a false-negative KRAS result (wild type versus G12C).Conclusions.-Molecular testing performed on H&E-stained sections via cell-transfer technique is useful when tissue from cell blocks and small surgical biopsy samples is exhausted and the only available material for testing is on H&E-stained slides.

Collection and Handling of Thoracic Small Biopsy and Cytology Specimens for Ancillary Studies: Guideline From the College of American Pathologists in Collaboration With the American College of Chest Physicians, Association for Molecular Pathology, American Society of Cytopathology, American Thoracic Society, Pulmonary Pathology Society, Papanicolaou Society of Cytopathology, Society of Interventional Radiology, and Society of Thoracic Radiology

“…Cytology laboratories use a wide variety of collection media, fixatives, and stains for specimen preparation. A wide range of cytology specimen processing techniques were identified in the systematic review, including CytoLyt (Hologic Inc, Marlborough, Massachusetts), [141][142][143] Hanks, 44 RPMI, 144 saline, 111,132,[145][146][147][148][149][150][151][152][153][154][155] and CytoRich (Becton, Dickinson and Company, Franklin Lakes, New Jersey) 43 as collection media; alcohol, 6,46,115,156 formaldehyde, 112 Mount-Quick, 119 and formalin || as fixatives; and Diff-Quik, 6,44,46,166 Giemsa, 43 and Papanicolaou for staining. 46,167 However, none of the listed studies performed direct comparisons between the different processing techniques that enable making a specific recommendation for the use of one method over another.…”

“…Although no study directly compared the use of one fixative with another, 4 studies 70,91,119,120 reporting on adequacy for ancillary testing or successful ancillary testing following fixation with 10% neutral buffered formalin comprise the indirect evidence supporting this statement. Studies reporting on mutational analysis comprise 2 prospective cohort studies, 70,119 and 2 retrospective cohort studies. 91,120 One of the prospective cohort studies 70 also reported on IHC and FISH, and 1 retrospective cohort study 91 reported on FISH in addition to mutational analysis.…”

“…91,120 One of the prospective cohort studies 70 also reported on IHC and FISH, and 1 retrospective cohort study 91 reported on FISH in addition to mutational analysis. Included studies carried an aggregate very serious risk of bias and were limited by their risk of bias in relation to patient selection, 70,91,119,120 detection, 91,120 and reporting, 91,120 as well as a lack of reported funding. 119,120 None of the studies were found to have methodologic flaws that would raise concerns about the findings.…”

“…Included studies carried an aggregate very serious risk of bias and were limited by their risk of bias in relation to patient selection, 70,91,119,120 detection, 91,120 and reporting, 91,120 as well as a lack of reported funding. 119,120 None of the studies were found to have methodologic flaws that would raise concerns about the findings. Refer to Supplemental Table 15 in the supplemental digital content for the quality assessment results of all studies included in the evidentiary base of statement 13.…”

Fine needle aspiration and core needle biopsy of metastatic malignancy of unknown primary site

“…Molecular testing performed on H&E-stained sections via CTT has also been utilized when tissue from cell blocks and small surgical biopsy samples were exhausted and the only available material for testing was present on the H&E-stained slides. Wu et al [36] applied PCR-based molecular testing (EGFR, BRAF, KRAS) using CTT on 97 samples, with 85% success rate, no significant residual tissue in the paraffin block for ancillary testing, with similar success.…”