MALDI imaging as a specific diagnostic tool for routine cervical cytology specimens

Cervical squamous cell carcinoma (SCC) is among the most common malignancies in women worldwide. In developed countries routine cytology screening has dramatically reduced SCC mortality within the last three decades. High risk (HR) human papilloma virus (HPV) infection is the main causal factor in nearly 100% of invasive SCCs, in most cases of low grade squamous intraepithelial lesion (LSIL) and in nearly all cases of high grade squamous intraepithelial lesion (HSIL). Detection of HR-HPV DNA has been extensively evaluated for the triage of patients with low grade cytological abnormalities in order to identify those at greatest risk for underlying or developing HSIL or SCC. However, the vast majority of HR-HPV-positive precursor lesions will not progress to invasive cancer. A variety of other screening tools are available which aim to stratify clinically significant HPV infections and cytological alterations. Matrix assisted laser desorption/ionization (MALDI) imaging mass spectrometry is a promising technology to assist in this endeavor. It delivers accurate mass spectrometric information of the sample's proteins and enables the visualization of the spatial distribution of protein expression profiles in correlation with histological features. In this study, 18 samples with Pap IIID or higher (>LSIL) and 14 samples with Pap I-II (WNL) were analyzed by MALDI imaging mass spectrometry (IMS). A genetic algorithm was applied to classify spectra resulting in an overall cross validation, sensitivity for Pap IIID and Pap?I-II of 83.7%, 88.9% and 78.6%, respectively. As this IMS based approach allows for unbiased and automated classifiction of cytological samples it appears to be a promising tool for stratification of cervical Pap smears.     

Quantitative histology as a diagnostic tool for celiac disease in children and adolescents

ObjectiveTo measure the villous height, the crypt depth, and the number of intraepithelial lymphocytes/100 enterocytes of the small intestinal mucosa of children and adolescents with celiac disease; and to classify these findings according to Q- Marsh and Q-histology scales.MethodsRetrospective study of a database from the Department of Pathology of biopsies from the second portion of the duodenum of pediatric patients. According to the histological report, three groups were established: celiac disease at diagnosis (n = 50), controls (n = 26), giardiasis (n = 10). In each biopsy, software (cellSens and Image J) evaluated 5 villous heights, 5 crypt depth and the number of intraepithelial lymphocytes/100 enterocytes.ResultsThe celiac group had the lowest mean villous height (197.83 μm) of all three groups (control = 477.70 μm; giardiasis = 397.04 μm. The celiac group's villous:crypt ratio (0.78) was significantly lower than the control group (1.89). The number of intraepithelial lymphocytes ≥25 was exclusive to the celiac group, with a sensitivity and specificity of 100 %. Only celiac patients were included in types 2 and 3 of the Q-histology classification.ConclusionCeliac disease patients showed shorter villous height than other groups, and the number of intraepithelial lymphocytes ≥25 was the best parameter to differentiate celiac from controls and giardiasis groups. Intraepithelial lymphocytes ≥25/100 enterocytes associated with any degree of villous atrophy, the classic Marsh 3 type, set the histological parameters of celiac disease. Quantitative histology is a valuable tool for diagnosing celiac disease, enabling histological changes in a short time, and the Q-histology scale appears to be more suitable than the Q-Marsh scale.     

Microarray-based genomic profiling as a diagnostic tool in acute lymphoblastic leukemia

In acute lymphoblastic leukemia (ALL) specific genomic abnormalities provide important clinical information. In most routine clinical diagnostic laboratories conventional karyotyping, in conjunction with targeted screens using e.g., fluorescence in situ hybridization (FISH), is currently considered as the gold standard to detect such aberrations. Conventional karyotyping, however, is limited in its resolution and yield, thus hampering the genetic diagnosis of ALL. We explored whether microarray-based genomic profiling would be feasible as an alternative strategy in a routine clinical diagnostic setting. To this end, we compared conventional karyotypes with microarray-deduced copy number aberration (CNA) karyotypes in 60 ALL cases. Microarray-based genomic profiling resulted in a CNA detection rate of 90%, whereas for conventional karyotyping this was 61%. In addition, many small (< 5 Mb) genetic lesions were encountered, frequently harboring clinically relevant ALL-related genes such as CDKN2A/B, ETV6, PAX5, and IKZF1. From our data we conclude that microarray-based genomic profiling serves as a robust tool in the genetic diagnosis of ALL, outreaching conventional karyotyping in CNA detection both in terms of sensitivity and specificity. We also propose a practical workflow for a comprehensive and objective interpretation of CNAs obtained through microarray-based genomic profiling, thereby facilitating its application in a routine clinical diagnostic setting.     

Volatile organic compounds in exhaled breath as a diagnostic tool for asthma in children

Background The correct diagnosis of asthma in young children is often hard to achieve, resulting in undertreatment of asthmatic children and overtreatment in transient wheezers.
Objectives To develop a new diagnostic tool that better discriminates between asthma and transient wheezing and that leads to a more accurate diagnosis and hence less undertreatment and overtreatment. A first stage in the development of such a tool is the ability to discriminate between asthmatic children and healthy controls. The integrative analysis of large numbers of volatile organic compounds (VOC) in exhaled breath has the potential to discriminate between various inflammatory conditions of the respiratory tract.
Methods Breath samples were obtained and analysed for VOC by gas chromatography–mass spectrometry from asthmatic children ( n =63) and healthy controls ( n =57). A total of 945 determined compounds were subjected to discriminant analysis to find those that could discriminate diseased from healthy children. A set of samples from both asthmatic and healthy children was selected to construct a model that was subsequently used to predict the asthma or the healthy status of a test group. In this way, the predictive value of the model could be tested.
Measurements and main results The discriminant analyses demonstrated that asthma and healthy groups are distinct from one another. A total of eight components discriminated between asthmatic and healthy children with a 92% correct classification, achieving a sensitivity of 89% and a specificity of 95%.
Conclusion The results show that a limited number of VOC in exhaled air can well be used to distinguish children with asthma from healthy children.
Cite this as : J. W. Dallinga, C. M. H. H. T. Robroeks, J. J. B. N. van Berkel, E. J. C. Moonen, R. W. L. Godschalk, Q. Jöbsis, E. Dompeling, E. F. M. Wouters and F. J. van Schooten, Clinical & Experimental Allergy , 2010 (40) 68–76.     

Fourier transform infrared microspectroscopy as a quantitative diagnostic tool for assignment of premalignancy grading in cervical neoplasia

The early diagnosis and proper identification of cervical squamous intraepithelial lesions plays an important role in a good prognosis for the patient. However, the present practice of screening based on PAP (Papanicolaou) smear and histopathology makes it tedious and prone to human errors. We assess the validity of FTIR microspectroscopy (FTIR-MSP) of biopsies as a method to properly assign the correct stage of premalignancy in patients with symptoms of cervical intraepithelial neoplasia. For the first time we evaluate the biopsies based on the FTIR spectra for different grades of neoplasia in tandem with probabilistic neural networks (PNNs) and histopathology. The results show that the grading of neoplasia based on FTIR-MSP and a PNN differentiates the normal from premalignant with a high level of accuracy. The false positive identification of the normal as cervical intraepithelial neoplasia 1 (CIN1), CIN2, and CIN3 patients is 9.04, 0.01, and 0.01%, respectively. The false negative identification of CIN2 patients as normal and CIN1 patients is 0.01 and 4.4%, respectively. Similarly, the false negative identification of CIN3 patients as normal, CIN1, and CIN2 is 0.14, 6.99, and 9.61%, respectively. The small errors encountered in the grading are comparable to current methods, encouraging advanced studies for the development of mechanized equipment for the diagnosis and grading of premalignant cervical neoplasia.     

Hair as a diagnostic tool in dysmorphology

Clinical diagnosis in dysmorphology is made by the recognition of a specific pattern of malformations and through an analytic search for discrete features.We present our personal experience regarding the usefulness of hair morphology as a tool for diagnosis in some metabolic and malformation syndromes. These cases represent only a few illustrative examples; an exhaustive review of the topic can be found elsewhere.     

Polymerase chain reaction as a rapid diagnostic tool for therapy of acute retinal necrosis syndrome

Herpesviruses are involved in the pathogenesis of acute retinal necrosis syndrome (ARN). A rapid and accurate diagnosis of herpetic infections is crucial for prompt administration of a specific antiviral therapy. The purpose of this study was to evaluate a polymerase chain reaction (PCR)-based assay to detect herpesvirus DNA in the aqueous humor of clinical samples from ten patients with uveitis and clinical suspicion of ARN. Samples were assayed for herpes simplex virus type 1-2 (HSV 1-2), varicella zoster virus (VZV) and cytomegalovirus (CMV). Clinical suspicion of ARN was confirmed for four patients. Two patients (one with bilateral ARN) tested PCR-positive for VZV DNA and the other two were positive for HSV 1-2 DNA. CMV DNA was not detected in any of the samples, and no sample was positive for DNA from more than one virus. The remaining patients did not show any evidence of herpesvirus DNA in their aqueous samples. Our findings demonstrate that the use of PCR for detecting herpesvirus DNA in aqueous humor of uveitic subjects may be a valuable tool for early diagnosis of acute retinal necrosis syndrome and for timely administration of a suitable therapy.     

Multifractal texture analysis of perfusion lung scans as a potential diagnostic tool for acute pulmonary embolism

A computer-assisted diagnostic (CAD) tool was developed for the diagnosis of acute pulmonary embolism (PE) in perfusion lung scans. Forty-five scans (with angiographic proof) were included in the study. The CAD tool was composed of two modules. The first module performs multifractal texture analysis on the posterior view of the perfusion scan. The second module is a decision algorithm that merges the multifractal parameters into a diagnosis regarding the presence or absence of PE. Linear and non-linear decision models were evaluated for the diagnostic task. A consensus neural network significantly outperformed all decision models including the physicians.     

Multispectral imaging as a tool for melanin detection

Multispectral imaging is becoming an increasingly useful tool in the field of histology. The ability to spectrally separate chromophores based on their differing spectra has been greatly beneficial when it comes to visualizing melanin in skin samples. Traditionally, Fontana Masson (FM) has been the gold standard for staining melanin, but with advances in technology, it is now possible to clearly detect and quantify melanin in hematoxylin and eosin (H&E) stained samples. With this advancement, there is no need to rely on a special stain to detect melanin in samples. Melanin detection and quantitation is critical for the study of hyperpigmentation and skin lightening. This study demonstrated that multispectral imaging is more sensitive than FM and can detect and quantify melanin that is co-localized in small quantities throughout skin samples.     

Rapid immunofiltration assay as a field diagnostic tool for ovine brucellosis

This work describes the development of two rapid immunofiltration assays, enzymatic (ERIFA) and non-enzymatic (NERIFA), for the rapid detection of ovine anti-Brucella antibodies. Brucella abortus lipopolysaccharide and total bacterial extract were dotted separately as diagnostic antigens on a nitrocellulose filter-membrane of the individual assay unit along with a third dot of purified sheep IgG as an internal control. The assay's diagnostic performance was evaluated in comparison with a modified rose bengal test (mRBT) and an indirect enzyme-linked immunosorbent assay (ELISA) through usage of 590 serum samples from healthy, vaccinated, or infected sheep. The ERIFA and indirect ELISA were found to be significantly more sensitive than NERIFA, while mRBT was determined to be statistically equivalent to NERIFA. A perfect agreement (κ?=?0.984) and a statistical equivalence to indirect ELISA suggest that the bi-antigenic ERIFA can be used as an "individual rapid ELISA" for screening ovine anti-Brucella antibody both in the field and in limited laboratory conditions.     

Clinical aspects of MR colonography as a diagnostic tool

Since first described in 1997, MR colonography (MRC) has since been labelled as a promising new, non-invasive technique for examining the colon. At present time, the examination is ready to be implemented as a supplement to incomplete colonoscopy or preoperative colonic evaluation. Furthermore, MRC seems to have a great potential in the screening for colorectal cancer, since detection of polyps and polypectomy might reduce on the incidence of colorectal cancer. This is speculated in the adenoma-carcinoma sequence theory, which states that most cancers evolve from polyps over a long period and that polypectomy might be curative. Colonoscopy remains the gold standard for full colon evaluation. However, the result of our studies can justify clinical use of MRC on selected indications, e.g. in the cases where colonoscopy is incomplete or technically difficult. Since up to 54% of all preoperative colon evaluations in patients with colorectal cancer and up to 17-23% of regular colonoscopies are incomplete, the clinical potential of MRC is evident. Furthermore, in our studies we have shown the insufficiency of preoperative colonic evaluation by CC. In addition, considering the invasiveness, the serious complications (perforation, bleeding, death) and the lack of patient acceptance in colonoscopy, the need for a safe, patient friendly alternative examination with high sensitivity, is clear. In conclusion, in the three studies that made up this PhD thesis, we have shown: that there are some flaws to the present gold standard of colonic evaluation; that there is an increased morbidity and mortality in the group of patients with missed SC; that patients have a preference for MRC and for fecal tagging compared to CC and bowel purgation and that there is a potential gain in doing preoperative colonic evaluation with MRC on all patients with rectal- or sigmoid colon cancer.     

Indirect immunofluorescence as a diagnostic tool for parvovirus infection of broiler chickens

Nuclear fluorescence was seen in the epithelial cells of the duodenum of broiler chicks infected experimentally at 1-day-old with a parvovirus of chicken origin. No antigenic relationship was detected by this method between chicken and goose parvoviruses.     

Switch of cadherin expression as a diagnostic tool for Leydig cell tumours

Leydig cell tumours comprise about 3% of all testicular tumours. The pathogenesis of Leydig cell tumours is still poorly understood. We investigated testis with Leydig cell hyperplasia and Leydig cell tumours for their expression pattern of P‐ and N‐cadherin. We could show a switch of expression of P‐ and N‐cadherin in Leydig cell hyperplasia and Leydig cell tumours in comparison with normal Leydig cells. Cadherins could be established as a new immunohistochemical marker for this testicular tumour entity; their possible role in tumour genesis will be discussed.     

Comparative genomic hybridisation as a supportive tool in diagnostic pathology

AIMS: Patients with multiple tumour localisations pose a particular problem to the pathologist when the traditional combination of clinical data, morphology, and immunohistochemistry does not provide conclusive evidence to differentiate between metastasis or second primary, or does not identify the primary location in cases of metastases and two primary tumours. Because this is crucial to decide on further treatment, molecular techniques are increasingly being used as ancillary tools. METHODS: The value of comparative genomic hybridisation (CGH) to differentiate between metastasis and second primary, or to identify the primary location in cases of metastases and two primary tumours was studied in seven patients. CGH is a cytogenetic technique that allows the analysis of genome wide amplifications, gains, and losses (deletions) in a tumour within a single experiment. The patterns of these chromosomal aberrations at the different tumour localisations were compared. RESULTS: In all seven cases, CGH patterns of gains and losses supported the differentiation between metastasis and second primary, or the identification of the primary location in cases of metastases and two primary tumours. CONCLUSION: The results illustrate the diagnostic value of CGH in patients with multiple tumours.     

Fine-needle aspiration cytology as a tool for the early detection of testicular relapse of acute lymphoblastic leukemia in children

Fine-needle aspiration cytology (FNAC) of the testis was done in 31 children (33 aspirates) with acute lymphoblastic leukemia (ALL) either as part of an end-therapy protocol (Group I, n = 20) or because of clinical suspicion of testicular relapse (Group II, n = 13). In Group I, none of the smears showed evidence of leukemic cells. However, two of these patients developed subsequent testicular relapse within 1 yr and were considered false-negative cases. Blast cells were present in nine patients of Group II; in the remaining four patients no neoplastic cells were observed in the smears and none experienced a relapse after a mean follow-up of 18 mo. Our findings indicate that FNAC can be a valuable method to evaluate clinically suspected testicular infiltration in children with ALL, and can be considered as an alternative procedure to surgical biopsy for screening testicular recurrence of childhood ALL. © 1994 Wiley-Liss, Inc.     

Radiohistology as a new diagnostic method for barium granuloma

Barium granulomas are rare complications of the barium enema. They pose diagnostic problems to the gastroenterologist, who may suspect a carcinoma, and to the pathologist, who may have difficulty in determining the precise nature of the foreign body. From four cases we suggest a simple and quick diagnostic method: paraffin-block roentgenography.