Braddock-Carey syndrome: A 21q22 contiguous gene syndrome encompassing RUNX1.

In 1994, Braddock and Carey first reported two unrelated girls with a new multiple malformation syndrome. The primary features included Pierre Robin sequence, persistent neonatal-onset thrombocytopenia, agenesis of the corpus callosum, a distinctive facies, enamel hypoplasia, and severe developmental delay. Since that time, there have been multiple other reported patients with a similar phenotype. In addition, several reports of thrombocytopenia and developmental delay have been documented in association with deletions in the Down syndrome critical region at 21q22. The similarity of the reported cases with deletions involving 21q22 with the clinical presentation of the two patients with Braddock-Carey syndrome resulted in a reinvestigation of the genetic etiology of these two patients 20 years after the original study. This investigation provides evidence that the etiology of this and other "Fanconi-like" disorders represent a newly recognized contiguous gene deletion syndrome involving 21q22 and specifically, the RUNX1 gene. © 2016 Wiley Periodicals, Inc.

Validation of MYC and BCL6 rapid break apart digital fluorescence in situ hybridization assays for clinical use.

OBJECTIVE: Detection of gene rearrangements in MYC (a family of regulator genes and proto-oncogenes) and human B-cell lymphoma 6 (BCL6) using fluorescence in situ hybridization (FISH) are important in the evaluation of lymphomas, in particular diffuse large B-cell lymphoma (DLBCL) and Burkitt lymphoma. Our current clinical MYC and BCL6 FISH workflow involves an overnight hybridization of probes with digital analysis using the GenASIs Scan and Analysis instrument (Applied Spectral Imaging). In order to improve assay turnaround time SureFISH probes were validated to reduce the hybridization time from 16 hours down to 1.5 hours. METHODS: Validation was a four-phase process involving initial development of the assays by testing new probes in a manual protocol, and cytogenetic studies to confirm the probe specificity, sensitivity, and localization. In the next phase, the assays were validated as a manual assay. The third phase involved development of the digital FISH assays by testing and optimizing the GenASIs Scan and Analysis instrument. In the final phase, the digital FISH assays were validated. RESULTS: Cytogenetic studies confirmed 100% probe sensitivity/specificity, and localization patterns. Negative reference range cutoffs calculated from 20 normal lymph nodes using the inverse of the beta cumulative probability density function (Excel BETAINV calculation) were 11% inclusive for both manual and digital MYC and BCL6 assays. There was 100% concordance between the manual and digital methods. The shortened hybridization time decreased the overall workflow time by 14.5 hours. CONCLUSIONS: This study validates the use of the SureFISH MYC and BCL6 probes on formalin fixed paraffin embedded (FFPE) tissue sections using a hybridization time of 1.5 hours that shortened the overall workflow by 14.5 hours. The process described also provides a standardized framework for validating digital FISH assays in the future.

Quantitative Imaging Analysis Fluorescence In Situ Hybridization Validation for Clinical HER2 Testing in Breast Cancer.

CONTEXT.­: Quantitative imaging is a promising tool that is gaining wide use across several areas of pathology. Although there has been increasing adoption of morphologic and immunohistochemical analysis, the adoption of evaluation of fluorescence in situ hybridization (FISH) on formalin-fixed, paraffin-embedded tissue has been limited because of complexity and lack of practice guidelines. OBJECTIVE.­: To perform human epidermal growth factor receptor 2 (HER2) FISH validation in breast carcinoma in accordance with the American Society of Clinical Oncology/College of American Pathologists (ASCO/CAP) 2018 guideline. DESIGN.­: Clinical validation of HER2 FISH was performed using the US Food and Drug Administration-approved dual-probe HER2 IQFISH (Dako, Carpinteria, California) with digital scanning performed on a PathFusion (Applied Spectral Imaging, Carlsbad, California) system. Validation parameters evaluated included z-stacking, classifier, accuracy, precision, software, and hardware settings. Finally, we evaluated the performance of digital enumeration on clinical samples in a real-world setting. RESULTS.­: The accuracy samples showed a final concordance of 95.3% to 100% across HER2 groups 1 to 5. During clinical implementation for HER2 groups 2, 3, and 4, we achieved a final concordance of 76% (95 of 125). Of these cases, only 8% (10 of 125) had discordances with clinical impact that could be identified algorithmically and triaged for manual review. CONCLUSIONS.­: Digital FISH enumeration is a useful tool to improve the efficacy of HER2 FISH enumeration and capture genetic heterogeneity across HER2 signals. Excluding cases with high background or poor image quality and manual review of cases with ASCO/CAP group discordances can further improve the efficiency of digital HER2 FISH enumeration.

Chromosomal loss of 3q26.3-3q26.32, involving a partial neuroligin 1 deletion, identified by genomic microarray in a child with microcephaly, seizure disorder, and severe intellectual disability.

Neuroligin 1 (NLGN1) is one of five members of the neuroligin gene family and may represent a candidate gene for neurological disorders, as members of this family are involved in formation and remodeling of central nervous system synapses. NLGN1 is expressed predominantly in the central nervous system, where it dimerizes and then binds with ß-neurexin to form a functional synapse. Mutations in neurexin 1 (NRXN1) as well as two other members of the neuroligin family, NLGN3 and NLGN4, have been associated with autism and mutations in NLGN4 have also been associated with intellectual disability, seizures, and EEG abnormalities. Genomic microarray is recommended for the detection of chromosomal gains or losses in patients with intellectual disability and multiple congenital anomalies. Results of uncertain significance are not uncommon. Parental studies can provide additional information by demonstrating that the imbalance is either de novo or inherited, and therefore is more or less likely to be causative of the clinical phenotype. However, the possibility that even inherited deletions and duplications may play a role in the phenotype of the proband cannot be excluded as many copy number variants associated with neurodevelopmental conditions show incomplete penetrance and may be inherited from an unaffected parent. Here, we report on a patient with a 2.2 Mb deletion at 3q26.3-3q26.32-encompassing the terminal end of NLGN1 and the entire NAALADL2 gene-detected by genomic microarray, and confirmed by FISH and real-time quantitative PCR. The same size deletion was subsequently found in her healthy, asymptomatic, adult mother.

Allele-specific PCR with competitive probe blocking for sensitive and specific detection of BRAF V600E in thyroid fine-needle aspiration specimens.

OBJECTIVE: To detect BRAF V600E mutation in thyroid fine-needle aspiration (FNA) slides and needle rinses (NR). STUDY DESIGN: Tumor-enriched DNA was extracted from FNA smears, formalin-fixed paraffin-embedded (FFPE) sections, or NR specimens from 37 patients with confirmed papillary thyroid carcinoma or benign findings. An allele-specific primer selectively amplified the 1799 T>A BRAF mutation while simultaneously blocking amplification of wild-type (WT) BRAF with an unlabeled probe during PCR. Mutation detection was accomplished by melting analysis of the probe. RESULTS: Allele-specific/blocking probe PCR confirmed the BRAF mutation status for 20 of 24 paired FNA/FFPE samples previously tested by fluorescent probe real-time PCR. For the other 4 cases, the sensitive PCR method detected the BRAF mutation in all paired FNA/FFPE samples. Previously, the mutation had been detected in only the FFPE samples. The BRAF mutation was also detected in some NR specimens. CONCLUSION: Treatment of patients with thyroid nodules is guided by FNA biopsy, which can be scantly cellular, necessitating a sensitive test that can detect low levels of BRAF V600E mutation in a WT background. We report increased detection of BRAF V600E in FNA specimens using allele-specific/blocking probe PCR, which has an analytical sensitivity of 0.01%.

Application of the Roche Cobas® HPV 4800 in Formalin-Fixed, Paraffin-Embedded Samples for Head and Neck Squamous Cell Carcinomas.

Testing for high risk human papillomavirus (HR-HPV) status is standard of care in squamous cell carcinomas of the oropharynx as well as cervical lymph node squamous cell carcinomas of unknown primary origin. DNA or RNA in-situ hybridization (ISH) and p16 immunohistochemistry, widely used currently for HPV detection are operator-dependent. In addition, DNA ISH has a relatively low sensitivity, and p16 is not entirely specific for HR-HPV infection. In this study, we examined the performance of the cobas® HPV genotyping assay in formalin-fixed, paraffin-embedded (FFPE) samples of head and neck squamous cell carcinoma. FFPE samples from head neck and other anatomic sites tested by ISH and p16 for HR-HPV at ARUP Laboratories were selected for this study. Samples were deparaffinized, stained and micro-dissected for tumor contents followed by tissue lysis, then tested with cobas® for HR-HPV. All the samples were also tested by HPV Linear Array for confirmation. All (N = 18) high risk HPV positive specimens tested by cobas® were confirmed as positive by the Linear Array test. All the specimens tested as negative by cobas® were tested as negative (N = 5) or positive only for low risk HPV (N = 3) by Linear Array, as cobas® only detects HR HPV. Limits of detection for HPV16 and 18 were established at 160-320 and 320-1600 copies, respectively. Our data suggest that cobas® HR-HPV genotyping is a viable option for detection of HR-HPV in formalin-fixed, paraffin-embedded samples from head and neck and other anatomic sites and has been validated for clinical use.

Enrichment and detection of rare alleles by means of snapback primers and rapid-cycle PCR.

BACKGROUND: Selective amplification of minority alleles is often necessary to detect cancer mutations in clinical samples. METHODS: Minor-allele enrichment and detection were performed with snapback primers in the presence of a saturating DNA dye within a closed tube. A 5' tail of nucleotides on 1 PCR primer hybridizes to the variable locus of its extension product to produce a hairpin that selectively enriches mismatched alleles. Genotyping performed after rapid-cycle PCR by melting of the secondary structure identifies different variants by the hairpin melting temperature (T(m)). Needle aspirates of thyroid tissue (n = 47) and paraffin-embedded biopsy samples (n = 44) were analyzed for BRAF (v-raf murine sarcoma viral oncogene homolog B1) variant p.V600E, and the results were compared with those for dual hybridization probe analysis. Needle aspirates of lung tumors (n = 8) were analyzed for EGFR [epidermal growth factor receptor (erythroblastic leukemia viral (v-erb-b) oncogene homolog, avian)] exon 19 in-frame deletions. RESULTS: Use of 18-s cycles and momentary extension times of "0 s" with rapid-cycle PCR increased the selective amplification of mismatched alleles. A low Mg(2+) concentration and a higher hairpin T(m) relative to the extension temperature also improved the detection limit of mismatched alleles. The detection limit was 0.1% for BRAF p.V600E and 0.02% for EGFR exon 19 in-frame deletions. Snapback and dual hybridization probe methods for allele quantification of the thyroid samples correlated well (R(2) = 0.93) with 2 more BRAF mutations (45 and 43, respectively, of 91 samples) detected after snapback enrichment. Different EGFR in-frame deletions in the lung samples produced different hairpin T(m)s. CONCLUSIONS: Use of snapback primers for enrichment and detection of minority alleles is simple, is inexpensive to perform, and can be completed in a closed tube in <25 min.

B-RAF V600E mutational analysis of fine needle aspirates correlates with diagnosis of thyroid nodules.

OBJECTIVE: A mutation of B-type RAF kinase (B-RAF) represents the most common genetic alteration in papillary thyroid cancer (PTC), possibly signifying a more aggressive biology. Fine needle aspiration (FNA) represents the most useful initial diagnostic tool of thyroid nodules. Molecular analysis of the mutation status of B-RAF in thyroid nodule FNAs may provide guidance for treatment planning. STUDY DESIGN: Cross-sectional study. SUBJECTS AND METHODS: A retrospective chart review was undertaken for clinically relevant data of papillary thyroid cancer (PTC), follicular variant of PTC (FV-PTC), and nonmalignant goiters. After blinded pathologic review, histologic and cytologic samples were analyzed by LightCycler PCR (LCPCR) with allele-specific fluorescent probe melting curve analysis (FMCA) for the V600E mutation of B-RAF. RESULTS: Of the 45 patient samples analyzed, B-RAF mutation was found to be significantly higher in papillary carcinomas when compared to follicular variant of papillary thyroid carcinomas (55.6% vs 14.3%, P = 0.05). Pathologic B-RAF mutational status significantly correlated with cytologic B-RAF mutational status (P < 0.0001), cytologic interpretation (P = 0.012), and histologic diagnosis (P = 0.011). CONCLUSIONS: Determination of B-RAF V600E mutation of thyroid nodule FNAs by LCPCR may be a useful tool to guide treatment planning. These data support investigating the utility of this molecular marker in a prospective manner.

An inv(16) in Ph-negative cells of a chronic myelogenous leukemia patient after imatinib treatment.

Secondary chromosomal changes are known to develop in Philadelphia chromosome-negative (Ph-) cells of chronic myelogenous leukemia (CML) patients after treatment with imatinib mesylate, an ABL kinase inhibitor. We report here a novel case of a pericentric inversion of chromosome 16 as the sole cytogenetic abnormality in Ph- cells after treatment of Ph+ CML with imatinib.

Subcutaneous tumor seeding following needle core biopsy of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the most common primary hepatic tumor and one of the most common cancers worldwide. At present, there are two widely used and accepted methods for obtaining diagnostic material for establishing the likelihood of malignancy in a hepatic mass, namely fine-needle aspiration (FNA) cytology and needle core biopsy (NCB). In recent years, however, tumor cell seeding along the needle tract has been shown to be a risk associated with using these procedures to obtain a pathologic diagnosis. We report a case of a patient who presented with a nodule in the anterior abdominal wall at the expected location of the previous NCB tract. FNA biopsy of the abdominal wall lesion confirmed the presence of malignant cells consistent with HCC. The finding of tumor seeding within a NCB tract raises the question of the role of NCB in the diagnostic workup of focal liver lesions.

Tumor cell nuclei extraction from paraffin-embedded lymphoid tissue for fluorescence in situ hybridization.

In this study the authors evaluated a technique for isolating intact tumor nuclei from paraffin-embedded lymphoma samples before performing FISH testing to detect the lymphoma-specific trans-location t(11;14) that defines mantle cell lymphoma. Well-characterized surgical pathology cases of mantle cell lymphoma were identified from pathology archives. Thin sections were cut from the paraffin-embedded tissue blocks. One section was stained using hematoxylin and eosin and an area composed exclusively of malignant cells was identified and marked on the slide. The corresponding area of the tissue block corresponding to this region underwent needle core biopsy, and the tissue was processed to isolate tumor cell nuclei and deposited onto a glass slide. The paired sample preparations underwent routine FISH testing for detection of the t(11;14)(q13;q32) chromosomal trans-location. DNA probe hybridization quality was compared between the tissue and isolated nuclei. Individual tumor cell nuclei were successfully extracted from each of the tissue blocks. The t(11;14) trans-location was detected by FISH in all of the samples diagnosed as mantle cell lymphoma. The hybridization signals found in the nuclei of extracted tumor cells were bright, planar, and easily identified. Detection of signal was superior to that on whole tissue samples, where signals often overlapped or were truncated. This technique produces intact nuclei for analysis, preserves the tissue block for additional studies, and allows sampling of a specific area of the tissue block. This approach may be particularly useful when the amount of diagnostic tissue is limited.

Utility of BRAF V600E mutation detection in cytologically indeterminate thyroid nodules.

BACKGROUND: Fine needle aspiration (FNA) is widely utilized for evaluation of patients with thyroid nodules. However, approximately 30% are indeterminate for malignancy. Recently, a mutation in the BRAF gene has been reported to be the most common genetic event in papillary thyroid carcinoma (PTC). In this retrospective study, we assessed the utility of BRAF V600E mutation detection for refining indeterminate preoperative cytologic diagnoses in patients with PTC. METHODS: Archival indeterminate thyroid FNAs and corresponding formalin-fixed, paraffin-embedded (FFPE) surgical samples with PTC were identified in our patient files. DNA extracted from slide scape lysates and 5 mum FFPE sections were evaluated for the BRAF V600E mutation using LightCycler PCR and fluorescent melting curve analysis (LCPCR). Amplification products that showed deviation from the wild-type genomic DNA melting peak, discordant FNA and FFPE matched pairs, and all benign control samples, underwent direct DNA sequencing. RESULTS: A total of 19 indeterminate thyroid FNAs demonstrating PTC on FFPE surgical samples were included in the study. Using BRAF mutation analysis, the preoperative diagnosis of PTC was confirmed in 3/19 (15.8%) FNA samples that could not be conclusively diagnosed on cytology alone. However, 9/19 (47.4%) FFPE tissue samples were positive for the V600E mutation. Of the discordant pairs, 5/6 FNAs contained less than 50% tumor cells. CONCLUSION: When used with indeterminate FNA samples, BRAF mutation analysis may be a useful adjunct technique for confirming the diagnosis of malignancy in an otherwise equivocal case. However, overall tumor cell content of some archival FNA smear slides is a limiting factor for mutation detection.

Validation of break-apart and fusion MYC probes using a digital fluorescence in situ hybridization capture and imaging system.

INTRODUCTION: Detection of MYC translocations using fluorescence in situ hybridization (FISH) is important in the evaluation of lymphomas, in particular, Burkitt lymphoma and diffuse large B-cell lymphoma. Our aim was to validate a digital FISH capture and imaging system for the detection of MYC 8q24 translocations using LSI-MYC (a break-apart probe) and MYC 8;14 translocation using IGH-MYC (a fusion probe). MATERIALS AND METHODS: LSI-MYC probe was evaluated using tissue sections from 35 patients. IGH-MYC probe was evaluated using tissue sections from forty patients. Sections were processed for FISH and analyzed using traditional methods. FISH slides were then analyzed using the GenASIs capture and analysis system. RESULTS: Results for LSI-MYC had a high degree of correlation between traditional method of FISH analysis and digital FISH analysis. Results for IGH-MYC had a 100% concordance between traditional method of FISH analysis and digital FISH analysis. CONCLUSION: Annotated whole slide images of H and E and FISH sections can be digitally aligned, so that areas of tumor within a section can be matched and evaluated with a greater degree of accuracy. Images can be archived permanently, providing a means for examining the results retrospectively. Digital FISH imaging of the MYC translocations provides a better diagnostic tool compared to traditional methods for evaluating lymphomas.

Cytologic features of nipple aspirate fluid using an automated non-invasive collection device: a prospective observational study.

BACKGROUND: Detection of cytologic atypia in nipple aspirate fluid (NAF) has been shown to be a predictor of risk for development of breast carcinoma. Manual collection of NAF for cytologic evaluation varies widely in terms of efficacy, ease of use, and patient acceptance. We investigated a new automated device for the non-invasive collection of NAF in the office setting. METHODS: A multi-center prospective observational clinical trial involving asymptomatic women designed to assess fluid production, adequacy, safety and patient acceptance of the HALO NAF Collection System (NeoMatrix, Irvine, CA). Cytologic evaluation of all NAF samples was performed using previously described classification categories. RESULTS: 500 healthy women were successfully enrolled. Thirty-eight percent (190/500) produced fluid and 187 were available for cytologic analysis. Cytologic classification of fluid producers showed 50% (93/187) Category 0 (insufficient cellular material), 38% (71/187) Category I (benign non-hyperplastic ductal epithelial cells), 10% (18/187) Category II (benign hyperplastic ductal epithelial cells), 3% (5/187) Category III (atypical ductal epithelial cells) and none were Category IV (unequivocal malignancy). Overall, 19% of the subjects produced NAF with adequate cellularity and 1% were found to have cytologic atypia. CONCLUSION: The HALO system is a simple, safe, rapid, automated method for standardized collection of NAF which is acceptable to patients. Cytologic assessment of HALO-collected NAF showed the ability to detect benign and pre-neoplastic ductal epithelial cells from asymptomatic volunteers.

Observations of the genomic landscape beyond 1p19q deletions and EGFR amplification in glioma.

BACKGROUND: With recent advancements in molecular techniques, the opportunities to gather whole genome information have increased, even in degraded samples such as FFPE tissues. As a result, a broader view of the genomic landscape of solid tumors may be explored. Whole genome copy number and loss of heterozygosity patterns can advance our understanding of mechanisms and complexity of various tumors. RESULTS: Genome-wide alterations involving copy number changes and loss of heterozygosity were identified in 17 glioma samples with positive FISH results for 1p19q co-deletions (n = 9) or EGFR amplification (n = 8). Gliomas positive for 1p19q co-deletions did not have other frequently recurrent genomic alterations. Additional copy-number alterations were observed in individual cases, and consisted primarily of large-scale changes, including gains or losses of entire chromosomes. The genomic architecture of EGFR amplified gliomas was much more complex, with a high number of gains and losses across the genome. Recurrent alterations in EGFR amplified gliomas were both focal, such as CDKN2A homozygous deletions, and large, such as chromosome 10 loss. CONCLUSIONS: Microarray enabled a broader picture of the genomic alterations occurring in glioma cases. Gliomas with 1p19q co-deletion had a relatively quiet genome, apart from the selected co-deletion. Additional alterations in isolated cases, involved primarily larger aberrations. On the other hand, EGFR amplified cases tended to be more complex and have specific abnormalities associated with the EGFR amplification. Furthermore, 1p19q co-deletions and EGFR amplification associated copy number changes appeared to often be mutually exclusive.

Streamlining the OncoScan® Array Procedure for Use in a Clinical Laboratory.

Microarray analysis has found tremendous utility in the clinical laboratory testing for detection of copy number changes (CNCs) and loss of heterozygosity (LOH). Recently the OncoScan® array was introduced as a tool for identification of CNCs and LOH in formalin-fixed paraffin-embedded oncology samples. The objective of this study was to identify steps in the OncoScan procedure that could be modified to make the process more efficient and technician-friendly in the clinical laboratory setting. Eighteen samples previously processed according to the manufacturer-recommended protocol were reprocessed using a modified protocol. The two primary modifications to the protocol included the elimination of a brief "chill and spin" step and an adjustment to the overnight hybridization temperature to allow for simultaneous hybridization of OncoScan and CytoScan® arrays. A comparison of paired samples processed using both protocols showed that our modified protocol performs similarly to the manufacturer-recommended protocol, yielding equivalent quality control metrics and calls.

The unsatisfactory ThinPrep Pap Test: missed opportunity for disease detection?

Cervical cytology specimens classified as "unsatisfactory for interpretation" represent a potential source of undetected disease. This prospective study analyzed the potential benefits of a laboratory procedure to reprocess unsatisfactory ThinPrep Pap Tests (Cytyc, Boxborough, MA). All unsatisfactory ThinPrep samples were reprocessed using a glacial acetic acid wash. The study period unsatisfactory rate was compared with that for the previous 12 months. The initial unsatisfactory rate was 1.3% (197/15,154). Of the unsatisfactory ThinPrep samples, 55.8% (110/197) had residual material for reprocessing. After reprocessing, 67.3% (74/110) were reclassified as "satisfactory" or "satisfactory but limited by," and the final unsatisfactory rate was 0.8% (123/15,154), a 62% decrease. Compared with the previous 12-month rate of 0.9% (209/23,730), this was a 12% reduction. Seven (6.4%) of 110 initially classified as unsatisfactory contained an epithelial abnormality (atypical squamous cells of undetermined significance, 3; atypical glandular cells of undetermined significance, 2; low-grade squamous intraepithelial lesion, 1; squamous cell carcinoma, 1) on the reprocessed slide. Reprocessing of unsatisfactory ThinPrep slides yielded additional cellular abnormalities that otherwise would have been undetected. The present study confirms that reprocessing of unsatisfactory ThinPrep slides is a beneficial laboratory procedure.

A stiff bristled, spiral-shaped ectocervical brush: a device for transepithelial tissue biopsy.

OBJECTIVE: To compare a new spiral-shaped tissue-sampling brush with a standard cervical punch biopsy. METHODS: Before large loop excision of the transformation zone, women with cervical intraepithelial neoplasia underwent a transepithelial brush biopsy of a portion of a colposcopically identified lesion, followed by a punch biopsy of the remaining portion. Brush biopsy samples were processed using liquid-based cytology and cell block techniques. Diagnoses were made using a consensus of three pathologists. Brush biopsy samples without basal cells were considered inadequate. The histological diagnosis was compared with the brush biopsy and punch biopsy samples. Patient-reported pain and physician-reported bleeding for punch and brush biopsies were compared. RESULTS: Fifty-two women were enrolled in the study; 47 successfully completed the study protocol. Eight brush biopsy specimens were inadequate. Thirty-nine women showed abnormal pathology (human papillomavirus/cervical intraepithelial neoplasia I or worse) on large loop excision of the transformation zone, and 32 women had high-grade (or worse) lesions. The punch biopsy correlated with high-grade disease in 53.1% of these women. The brush biopsy result correlated with high-grade disease in 79.3% of these women using a cell block technique and 76.7% using liquid cytology. There was significantly less pain (P <.001) and significantly less bleeding (P <.001) with the brush biopsy. CONCLUSION: When an adequate sample is collected, spiral brush biopsy is as good as a standard punch biopsy for detecting cervical pathology, with substantially less pain and bleeding. User training and guidelines for sampling are needed to assure that an adequate sample is collected.

A simple method to determine the need for glacial acetic acid treatment of bloody ThinPrep Pap tests before slide processing.

ThinPrep (TP) Papanicolaou (Pap) samples containing excessive blood often result in unsatisfactory preparations, possibly leading to undetected gynecologic disease, and added inconvenience to patients and clinicians. Reprocessing of these samples with a glacial acetic acid wash is effective at eliminating blood, providing satisfactory preparation and detection of lesions. However, it increases laboratory costs and decreases work flow efficiency. We report the use of a color standard for gauging the necessity of performing a glacial acetic acid wash before TP processing. This "preprocessing" was found to reduce the costs associated with reprocessing by 48%, while maintaining high preparation quality by improved sample adequacy.

Prevalence and typing of HPV DNA by hybrid capture II in women with ASCUS, ASC-H, LSIL, and AGC on ThinPrep Pap tests.

Testing for human papillomavirus (HPV) DNA is now a viable option for the management of women with atypical squamous cells of undetermined significance (ASCUS). The utility of reflexive HPV DNA testing for women with a cytologic diagnosis of atypical glandular cells-not otherwise specified (AGC-NOS), ASCUS subtypes, and low-grade squamous intraepithelial lesion (LSIL) has not been well established. In the present investigation, reflex Hybrid Capture II HPV DNA testing results were evaluated for HPV prevalence and type in 371 women with abnormal cytologic diagnoses of ASCUS-not otherwise specified (ASCUS-NOS), ASCUS-suspicious for low-grade squamous intraepithelial lesion (ASCUS-L), atypical squamous cells-cannot exclude high-grade squamous intraepithelial lesion (ASC-H), AGC-NOS, and LSIL on ThinPrep Pap tests. Positive high-risk HPV DNA was identified in 53.6% of the study samples, including ASCUS-NOS 40.2% ASCUS-L 71.4%, ASC-H 37.5%, LSIL 88.6%, and AGC-NOS 0%. We conclude that reflex HPV DNA testing appears to not be useful for colposcopy triage for cytologic diagnoses of LSIL or AGC-NOS.