Sampling, sampling errors and specimen preparation.

To obtain an adequate cervical smear for making a correct cytologic diagnosis, smear taking, laboratory handling and interpretation must be optimal. Many people are involved, and only by a combined effort of all links can this target be seriously approached: the smear takers will have to be open minded about technical improvements and read the morphologic descriptions cautiously; in the laboratory, cytotechnicians and physicians will have to challenge themselves and each other. It is mandatory to discard specimens that do not meet general standards of adequacy. At present a host of new techniques are being implemented. It is not feasible for all laboratories to be engaged in testing these new methods, but we are all requested to follow the development the best we can and switch to new ways when justified. Our working conditions are very different; therefore, it is our professional responsibility and plight to respond at the right time. So far the conclusion is that the conventional Pap smear is the international standard of care for the diagnosis of cervical cancer precursers in cancer screening programs. Certainly, this may change within a very short time. Liquid-based techniques, and in particular HPV technologies, are just around the corner.

New technologies in gynecologic cytology.

New technologies are trying to make significant changes in the way we perform the Pap smear test, a test that has undergone no significant modification since its inception. Efforts have focused on improving sampling with liquid-based methods, improving detection of abnormal cells with automated screening and improving the detection of human papillomavirus with assays practical for use in routine clinical material. This article reviews the performance of the new technologies to date, with an emphasis on those that are currently approved for cervical cytology testing. In the aggregate, these technologies can have a positive impact on cervical disease detection and intervention.

Quality assurance and risk reduction guidelines.

Cervical cancer continues to be a major cause of death in women worldwide. The major problem facing most women is the unavailability of screening Pap tests in poor and underdeveloped countries. While rates of cancer deaths have decreased 60-80% in developed countries since the Pap test became available, the accuracy of Paps was challenged recently. In order to instill public confidence and promote optimal patient care, measures to improve the quality of the entire screening process should be undertaken. Continuous quality improvement processes are more appropriate than traditional quality assurance monitors. Although no standards can be defined that are applicable to all laboratory settings and nations, this document provides current views on universal quality procedures and risk reduction. Procedure/policy manuals, workload assessment, hierarchic/peer review, discrepancy analysis, rescreening studies and cytohistologic correlation are examples of universally applicable quality tools. The variability in practices in different parts of the world is also discussed.

AutoCyte Prep system vs. conventional cervical cytology. Comparison based on 2,156 cases.

OBJECTIVE: To compare the AutoCyte Prep system (Burlington, North Carolina) with conventional cervical cytology in a university medical center laboratory. STUDY DESIGN: Split-sample conventional and AutoCyte Preps were examined for 2,156 cases. Same-patient conventional and Prep slides were submitted to separate cytotechnologists blindly. The results were compared on review by a cytopathologist. The Prep slides were subsequently scanned on the AutoCyte Screen automated interactive system, with manual review of the flagged cases. The results were compared with anatomic pathology follow-up when available. RESULTS: Of the 158 squamous intraepithelial lesions (SILs) found by both methods, 78% were found by AutoCyte Prep, while 59% were found by conventional smear (P < .01). Among the discordant cases, thin-layer slides revealed 88% more LSIL lesions (P < .05), a comparable number of high grade SIL lesions and a single case of adenocarcinoma that was ambiguous on the conventional slide. The addition of AutoCyte assisted primary screening demonstrated a net benefit, recovering additional cases referenced as atypical glandular cells of undetermined significance and as high grade SIL. CONCLUSION: The AutoCyte Prep system affords excellent cellular presentations and superior sensitivity for SILs when compared to the conventional technique. The use of AutoCyte Screen, for primary screening demonstrated performance equivalent to manual screening, with a significant improvement in sensitivity when measured against biopsy results.

Evaluation of the AutoCyte SCREEN system in a clinical cytopathology laboratory.

OBJECTIVE: To compare the AutoCyte SCREEN (AutoCyte, Burlington, North Carolina, U.S.A.) system with manual screening by experienced cytotechnologists using thin-layer preparations that had been previously extensively studied and their cytologic abnormalities well defined. STUDY DESIGN: AutoCyte PREP (AutoCyte) samples prepared for a previous split-sample study comparing thin-layer preparations to conventional smears were used. These 1,992 AutoCyte PREP samples were in a cohort the abnormal findings of which had been confirmed via independent review by two sets of pathologists. For the current study, these samples were remasked and evaluated by the AutoCyte SCREEN system in a clinical laboratory. The instrument scanned each slide and selected six overview fields and 120 single objects for storage and display. The computer classified each slide in one of the following categories: abnormal, uncertain, normal or unsatisfactory. Independently for each case, a cytotechnologist evaluated the six fields and 120 objects selected by the instrument as abnormal, normal or unsatisfactory. For those cases classified as uncertain by AutoCyte, the technologist then reexamined the cellular displays and entered a consensus classification. These results were then compared to those of an independent review by cytotechnologists of the identical set of slides using routine manual screening. RESULTS: The AutoCyte SCREEN selected 35% of slides for manual review. Technologist and computer rendered equivalent classifications in 79%. Of the total slides screened by the AutoCyte SCREEN, 57% were classified as "uncertain," and 88% of these were subsequently classified as normal by consensus. Using the well-defined abnormal values of the cellular sample as a basis for calculation, the AutoCyte SCREEN-assisted practice had a diagnostic sensitivity of 85% and diagnostic specificity of 97.6%. Comparable values for manual screening of the identical cellular sample were a diagnostic sensitivity of 80% and specificity of 97.4%. CONCLUSION: The AutoCyte SCREEN achieves comparable or greater sensitivity in detecting cervical abnormalities in comparison with manual screening. When combined with the substantial advantage of thin-layer preparations over conventional smears, the AutoCyte SCREEN provides a screening system of superior sensitivity over conventionally prepared and examined cervical smears.

Multicenter comparison of manual and automated screening of AutoCyte gynecologic preparations.

OBJECTIVE: To compare AutoCyte SCREEN-assisted evaluation of AutoCyte PREP liquid-based preparations with manual microscopic screening of the same preparations in a masked, multisite trial. STUDY DESIGN: AutoCyte PREPs were made using the CytoRich automated, liquidbased method from the residual cellular material on the collection device after a conventional cervical smear had been made. The study involved 1,676 samples collected sequentially from high-risk patients at two medical centers. The AutoCyte PREPs were then screened manually by cytotechnologists at one of two laboratory sites. All abnormal slides were reviewed by the site pathologists for final diagnosis. The PREPs were then remasked and screened using the AutoCyte SCREEN automated, interactive screening system, designed to select potentially abnormal slides for manual review while allowing the direct sign-out of negative slides. The AutoCyte SCREEN-assisted practice result was determined by combining the interactive SCREEN result with manual evaluation for those cases selected by SCREEN for manual review. All slides deemed abnormal were manually reviewed by an independent reference pathologist. The original manual review results were then compared to the AutoCyte SCREEN-assisted practice results stratified by the Bethesda categories of abnormal diagnoses as determined by the reference pathologist. RESULTS: Of the 1,676 cases, 494 were determined to be abnormal (ASCUS+) by one or both of the study methods and also by the independent reference pathologist. Of these 494 abnormal cases, 312 had a reference diagnosis of LSIL+, and 139 had a reference diagnosis of HSIL or cancer. The remainder of these cases were ASCUS or AGUS. Sensitivities and false negative proportions were stratified by the reference pathologist based on Bethesda categories as "truth" and compared. For LSIL+ cases, manual screening alone had a sensitivity of 89% as compared to 98% for the AutoCyte SCREEN-assisted practice. Manual screening demonstrated 90% sensitivity to HSIL or greater abnormality as compared to 99% sensitivity by the AutoCyte SCREEN-assisted practice. CONCLUSION: There was a concurrent significant reduction in the false negative fraction using the AutoCyte SCREEN as part of screening practice. Specificity for both screening practices was equivalent.

Utility of residual AutoCyte cervical cytology samples for image analysis.

OBJECTIVE: To ascertain the utility of residual liquid-preserved cervical cytology samples for DNA profiling studies. STUDY DESIGN: Ninety-nine liquid-preserved cervical cytology samples from a high-risk population were received two months after the initial diagnosis at another laboratory. Each residual sample was processed to yield one Papanicolaou-stained and one Feulgen-thionin-stained slide. The former were examined manually without knowledge of the submitted findings and were also examined on the AutoCyte SCREEN interactive system. DNA profiling of > 185 cells per case was performed on an RPW image analyzer. Results were compared with submitted diagnoses. RESULTS: The 99 cases included 59 normals, 9 ASCUS/AGCUS, 16 HSIL, 12 carcinomas and 3 unsatisfactory. Interlaboratory agreement between residual and initial samples was good (unweighted kappa = .609). Abnormals were characterized by 2C deviation indices > 2.0 and 5C exceeding rates > 4.37%. AutoCyte SCREEN system examination of residual slides of high grade abnormalities was 100% sensitive and 40% specific. CONCLUSION: Residual AutoCyte cervical cytology samples are stable and yield reproducible results for routine and ancillary studies of cervical cytologic abnormalities. The AutoCyte SCREEN system was 100% sensitive for high grade abnormalities in this enriched sample, even when operating on residual material.

Disseminated mucosal papilloma/condyloma secondary to human papillomavirus.

This report details the histopathologic findings in a woman who acquired the human papillomavirus 6/11 in her late teens and developed papilloma/condyloma of the nasopharynx, oropharynx, anogenital region, urethra, and urinary bladder. General evaluations of immune function reveal no defect, and there was no evidence of HIV infection. The morphologic expression of HPV 6/11 infection appears to be completely dependent on the mucosal epithelium affected. The complete spectrum of benign and premalignant epithelial changes induced by the human papillomavirus family-papilloma, verrucae, condyloma acuminatum, epithelial hyperplasia, and dysplasia-were present in this patient with a single papillomavirus infection. We postulate that this patient has a specific immune deficiency that limits her ability to control local infection and spread of the papillomavirus.

Cell recovery and appearance in thin-layer preparations in nongynecologic cytology.

OBJECTIVE: To compare the diagnostic efficacy of a thin-layer technique with that of a membrane filter technique using a wide variety of fresh cytologic specimens. STUDY DESIGN: Paired samples from 272 nongynecologic cytology specimens were processed for microscopy using thin-layer and membrane filter preparation techniques. Specimens included 162 body fluids and urines, 32 bronchial aspirates and 78 fine needle aspiration biopsies. The two techniques were compared for diagnostic efficacy, cell density/specimen, cell types recovered, qualitative cytomorphology, ease of interpretation and long-term stability of the final preparations. RESULTS: Diagnoses supported by filter preparations were also supported by their matched thin-layer preparations. The same cell types were recovered by both techniques, and the slide cell density was slightly greater using membrane filters. Qualitative morphology and ease of use were superior with the thin-layer samples. After two years, all thin-layer slides were stable and readable, whereas only 43% of the membrane filter slides were usable. CONCLUSION: Thin-layer techniques are morphologically comparable and, in some ways, superior to membrane filter techniques for processing a wide variety of nongynecologic cytology specimens.

Cellular morphometry in nongynecologic thin-layer and filter cytologic specimens.

OBJECTIVE: To determine the feasibility and utility of thin-layer cytology preparations for morphometric analysis of nuclear and cytoplasmic area in cells from nongynecologic cytology specimens. STUDY DESIGN: Identical paired samples from nongynecologic specimens (bronchial aspirate, urine, pleural and peritoneal fluid) were used to prepare thin-layer preparations and corresponding traditional membrane filter preparations. The paired preparations were analyzed by static image morphometry measuring eight nuclear and cytoplasmic parameters that allowed calculation of nuclear area, cytoplasmic area and total cell size. Hepatocytes and bronchial columnar, mesothelial, squamous and transitional cells were studied, as was a single case of high grade transitional cell carcinoma. Sufficient numbers of each cell type were measured to allow statistical analysis. RESULTS: Both thin-layer and membrane filter preparation techniques yielded individual cells suitable for morphometric analysis, and there were no consistent morphologic measurement differences between the two methods. The thin-layer preparation had the following significant technical advantages: more numerous easily measured single cells, lack of interfering background and superior specimen stability. Benign oval to round nuclei from a variety of cell types have a mean nuclear area in a narrow range from 29 to 55 microns 2. The mean nuclear area of malignant cells studied was significantly larger (78 microns 2), and there was a significant decrease in absolute cytoplasmic profile area in the malignant population studied. CONCLUSION: Thin-layer cytology preparations have significant advantages for morphometric studies over traditional membrane filter cytologic preparations. The morphometric measurement of nuclear area in benign and malignant cell populations has great potential as a generic screening tool for malignancy in cytologic specimens. Simultaneous measurement of cytoplasmic area adds a powerful dimension. The subsequent calculation of a true nuclear/cytoplasmic ratio may yield a sensitive and specific discriminator for detecting malignant cell populations in human nongynecologic cytology specimens.

Performance and retention of professional employees who work in teams: the effects of commitment and support.

The purpose of this study was to determine the impact of support and commitment on job performance and turnover intentions of professionals working in a team environment. As hypothesized, the results indicated that job performance was influenced positively by both professional and team commitment as well as by perceived organizational and team support. Intention to quit the organization went down as organizational support and commitment went up, and intention to leave the profession similarly was related to team support and commitment Interestingly, professional commitment had a deleterious effect on intention to quit the organization, whereas organizational support had a positive effect on professional commitment.

Multicenter masked evaluation of AutoCyte PREP thin layers with matched conventional smears. Including initial biopsy results.

OBJECTIVE: To compare the performance of liquid-based cell preparations from the AutoCyte PREP system (AutoCyte, Inc., Elon College, North Carolina) with the conventional cervical smear in masked, split-sample, multisite trials. STUDY DESIGN: The AutoCyte PREP system utilizes the CytoRich method, which combines liquid preservation, selective reduction of blood and inflammation, thin-layer cell dispersion and discrete staining. In an eight-site multicenter trial, 8,983 cases were evaluated. Parallel AutoCyte PREP slides and matching conventional cervical cytologic smears were screened in a masked fashion, with all abnormals reviewed in a masked, on-site pathologist review. The conventional smear was always prepared first, with the AutoCyte PREP using the residual cells on the collection device. The Bethesda System was used for reporting diagnosis and specimen adequacy. RESULTS: Of the cases, 7,805 (86.9%) had the same interpretation. In 8,750 cases (97.4%), there was agreement within one diagnostic category. The AutoCyte PREP demonstrated a statistically significant, 31% overall improvement in the detection of squamous intraepithelial lesion and invasive cancer when evaluating cases with more than one diagnostic class difference. Biopsy correlation of available data supported the improvement observed in the cytology-based comparison. Recovery of infectious organisms and endocervical component was comparable between preparations, especially in light of the split-sample study design. There was a 39% reduction in unsatisfactory slides and 44% fewer satisfactory but limited by reports. CONCLUSION: The AutoCyte PREP and CytoRich method produce excellent cellular presentations with standardized quality, superior sensitivity and improved adequacy as compared to the conventional method.

Quantification of tissue eosinophils and lymphocytes in histologic sections.

During a study of eosinophil-predominant gallbladder disease, an image analysis (IA) technique was developed for quantification of eosinophils and lymphocytes in routine formalin-fixed tissue sections. Alternating sections were stained with hematoxylin and eosin for eosinophils and with monoclonal CD45 antibody visualized with diaminobenzidine by an avidin-biotin procedure for lymphocytes. A protocol was then developed using a commercially available image analyzer and two well-defined macro routines. The system was validated with cell block sections prepared from peripheral blood samples with known eosinophil and lymphocyte counts. The eosinophil counts obtained by this IA technique showed excellent correlation with the absolute counts from a peripheral blood analyzer (r2 = 0.987). The lymphocyte counts obtained by IA showed good correlation with the absolute counts (r2 = 0.820). This IA-based technique provides a sensitive, reproducible, and substantiated means of quantifying inflammatory cells in tissue sections. This rapid and easily learned technique adds quantification as a complementary dimension to the subjective assessment of tissue morphology.

The cost of production in cervical cytology: comparison of conventional and automated primary screening systems.

To thrive in a world of managed care and capitation, knowing the cost of production is critical, especially in cervical cytology. The relative costs of production for preparation and interpretation of cervical cytology for conventional manually read smears and a primary automated screening method were calculated in a university practice setting. The components were disposables, processing, screening, pathologist review, capital equipment, and facilities cost. The production cost for a conventional smear in our laboratory is $9.75, and the comparable production cost by a primary screening automated method would be $12.07, if it were approved for primary screening. Based on these calculations, an interactive automated method approved for primary screening would cost slightly more than conventional testing. Combinations of conventional and automated examinations (secondary screening systems) are not cost feasible unless the additional production expense can be passed through to the payers.

The cost of diagnosis: a comparison of four different strategies in the workup of solitary radiographic lung lesions.

INTRODUCTION: Due to the large numbers of negative results of sputum examinations in the face of malignant disease, we used a decision analytic model to determine whether, and under what conditions, sputum cytology (Spt) might have a cost-effective role to play in the approach to lung lesions. METHODS: We constructed a decision analytic model to compare the utility of Spt, image-directed fine-needle aspiration (FNA), bronchoscopic examination (Bronch), and open biopsy (OBx) in the evaluation of lung lesions. Prevalence and cost data were derived from local databases and diagnosis-related groups. Diagnostic sensitivity (sens) and specificity (spec) of the tests were derived from the literature and local data. Output of the model was lowest cost per correct malignant diagnosis and included surgical treatment costs. We did not attempt to model survival data or morbidity. Sensitivity analyses were performed using cost, test sensitivity, and lesion size variables. RESULTS: In the baseline case, a patient who is a surgical candidate with lesion size 2.8 cm, prevalence of malignancy=0.67, FNA sens=0.95, Bronch sens=0.80, Spt sens=0.51, OBx is the best initial procedure with a cost per correct diagnosis of $12,888. Sputum examination has the highest cost per correct diagnosis of $63,424. FNA and bronchoscopy have cost per correct diagnosis of $21,543 and $16,615, respectively. Sens analysis shows that OBx is the preferred strategy in the workup of lung lesions in patients who are surgical candidates older than 30 years of age. Spt is the preferred strategy only when the patient is not a surgical candidate, the lesion size is large (>4.7 cm), and only if sputum sens is >0.45. CONCLUSIONS: Spt is not cost-effective in clinical practice, except when the patient has a large clinically unresectable lesion and the laboratory can demonstrate sufficient test sens.

Time interval effect on repeat cervical smear results.

OBJECTIVE: Second cervical smears obtained at short time intervals often exhibit a lesser degree of abnormality than the first smear. We studied the effect of time interval between smears on diagnoses in two large, distinctive cohorts. STUDY DESIGN: Patients with two or more satisfactory smears with at least one smear or a cervical biopsy showing atypical squamous cells of undetermined significance or greater were selected. Patients were divided into four subsets by test intervals (days) (< or = 45, 46-90, 91-120, > 120) and compared statistically. RESULTS: The distribution of differences between results for the short-interval subsets (< 120) was significantly different (P < .01) from the interval subset > 120 days. At short intervals the results revealed loss of sensitivity in the second smear as compared to the initial smear and concurrent biopsies. CONCLUSION: Rapidly repeated cervical smears show poor agreement with the biopsy and may be misleading. This effect is most pronounced when the interval is < 45 days. Colposcopists should consider whether concurrent smears shortly after an abnormal smear are worth performing, given the loss of sensitivity.

Comparison of the CytoRich system with conventional cervical cytology. Preliminary data on 2,032 cases from a clinical trial site.

OBJECTIVE: To compare the CytoRich system with conventional cervical cytology in a university medical center hospital laboratory. STUDY DESIGN: The CytoRich system combines liquid preservation, selective reduction of blood/inflammation, thin-layer preparation and discrete staining. Two thousand thirty-two parallel conventional and CytoRich samples were examined as part of a multicenter trial of the CytoRich/AutoCyte systems. Same-patient conventional and CytoRich slides were submitted to separate cytotechnologists blindly. The results were compared, and all nonmatching sample pairs were reviewed again. A consensus diagnosis was derived for all cases. The initial readings of the CytoRich and conventional smears were compared with each other and with the consensus diagnosis. RESULTS: Of the 148 squamous intraepithelial lesions (SILs) found by either method, 85% were found by CytoRich, while only 58.5% were found by conventional smear. As compared with the consensus diagnosis, CytoRich slides had 86.7% sensitivity for SIL and 99.1% specificity, while the conventional slides had 63.6% sensitivity and 99.7% specificity. Consensus review resulted in upgrading to SIL in 1.8% of conventional slides and 1.4% of CytoRich slides. The biopsy correlation results were similar for the two methods. CONCLUSION: The CytoRich system affords excellent cellular presentations and superior sensitivity for SILs when compared to the conventional technique.

Evaluation of red cell volume distribution width (RDW).

The purpose of this study was to evaluate red cell distribution width (RDW) in the differential diagnosis of patients with well defined iron deficiency (ID), anemia of chronic disease (ACD), and thalassemia trait. Mean RDW in 100 "normals" was 13.1%. Elevated RDW (greater than 15.0%) was observed in 90% of ID (N = 39), 32% of ACD(N = 79), 66% of beta-thalassemia trait (N = 100), 53% of alpha-thalassemia trait men (N = 127), 34% of alpha-thalassemia trait women (N = 116), 3% of sickle cell trait (N = 101), and 88% of sickling disorders (N = 32). The Bessman classification by mean corpuscular volume and RDW was of limited value in separating ID from thalassemia trait but was improved by adopting 16.0% as the upper limit of normal for RDW. Although many attempts have been made to discriminate between ID and thalassemia trait on the basis of red cell values from a cell counter, none (including RDW) is sufficiently specific or sensitive to obviate the need for confirmation of the diagnosis by standard laboratory methods.

Multiple hormone production in an oat cell carcinoma of the larynx.

A 60-year-old white woman with laryngeal oat cell carcinoma is described. She was a heavy smoker who had been treated seven years earlier with 5,000 rads for a well differentiated squamous cell carcinoma metastatic to a left submandibular lymph node. She presented this time with a two month history of hoarseness and tumor of the supraglottic larynx was found. There was clinical and chemical evidence of an ectopic ACTH syndrome. The histology and fine structure of the tumor were typical of oat cell carcinoma. Immunoreactive ACTH, GRP, NSE, Beta-endorphin, calcitonin, and keratin were found in the cytoplasm of the tumor cells by indirect immunoperoxidase techniques. We could find no previously reported case of laryngeal oat cell carcinoma with ectopic ACTH syndrome or cytoplasmic localization of polypeptides.