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PubMed2001

Determination of diamino acids in peptidoglycans from anaerobic bacteria.

Peptidoglycans isolated from two Fusobacterium species of anaerobic bacteria were analyzed for constituent amino acids. Hydrolysis conditions were varied to optimize the yield of diamino acids from peptidoglycan. The o-phthalaldehyde derivatives of the diamino acid stereoisomers were separated by high-performance liquid chromatography (OPA-HPLC), and variations in the relative areas of the two peaks noticed during analysis of solid samples were attributed to sampling errors. Co-derivatization/injection experiments using standards of the meso and rac forms separated from commercial mixtures demonstrated that meso-2,6-diaminopimelic acid and meso-lanthionine were peptidoglycan components in Fusobacterium varium and Fusobacterium nucleatum, respectively. The protonated molecules of 2,6-diaminopimelic acid and lanthionine were detected in peptidoglycan hydrolyzates by off-line, flow-injection electrospray mass spectrometry (ESI-MS). In ESI-MS-MS experiments under identical collision-induced dissociation (CID) conditions, peptidoglycan-derived and standard diamino acids exhibited similar fragmentations. Fragmentation pathways are proposed for each diamino acid. The results confirm that the meso forms of two different diamino acids are utilized in the peptidoglycans of Fusobacterium species.

Amino Acids↗
PubMed2002

[Breast calcifications with percutaneous vacuum-assisted biopsy diagnosis of malignancy or atypical hyerplasia: correlations with surgical findings].

Percutaneous, stereotactic, vacuum-assisted biopsy has become a widely used alternative to open surgical biopsy for the initial diagnosis of breast calcifications. We retrospectively assessed the accuracy of the technique in the diagnoses of malignancy and atypical hyperplasia by correlation with the findings of the subsequent surgical excision. We studied 330 consecutive cases of breast calcifications, 216 (65.5%) of which were determined to be benign and 114 (34.5%) to be malignant or atypical at vacuum-assisted biopsy using an 11 gauge instrument. Of the latter 93 were available for comparison with the subsequent surgery, the specific diagnoses as revealed by percutaneous biopsy were as follows: 11 cases of atypical ductal hyperplasia (ADN), 67 cases of ductal carcinoma in situ (DCIS), 6 infiltrating ductal carcinomas (IFDC), 2 cases of atypical lobular hyperplasia and 7 of lobular carcinoma in situ (LCIS). At histological analysis after surgical excision, 3 (27%) of 11 cases previously diagnosed as ADH and 6 (9%) of 67 cases diagnosed as DCIS were shown to actually be higher grade lesions (DCIS/IFDC and IFDC, respectively). Of the 7 lesions diagnosed at vacuum-assisted biopsy as LCIS, surgery and histological analysis showed one infiltrating globular carcinoma and two DCIS. A total of 21 lesions (4 ADH, 14 DCIS, 1 IFDC, 2 LCIS) were completely removed at percutaneous biopsy; the remaining cases were found totally concordant. These data Indicate a substantial accuracy of the percutaneous biopsy: some lesions (particularly those thought to be ADH and DCIS) can be underestimated for sampling error.

Biopsy, Needle↗
PubMed2005

Cholesky problems.

Behavioral geneticists commonly parameterize a genetic or environmental covariance matrix as the product of a lower diagonal matrix postmultiplied by its transpose-a technique commonly referred to as "fitting a Cholesky." Here, simulations demonstrate that this procedure is sometimes valid, but at other times: (1) may not produce fit statistics that are distributed as a chi2; or (2) if the distribution of the fit statistic is chi2, then the degrees of freedom (df) are not always the difference between the number of parameters in the general model less the number of parameters in a constrained model. It is hypothesized that the problem is related to the fact that the Cholesky parameterization requires that the covariance matrix formed by the product be either positive definite or singular. Even though a population covariance matrix may be positive definite, the combination of sampling error and the derived--as opposed to directly observed--nature of genetic and environmental matrices allow matrices that are negative (semi) definite. When this occurs, fitting a Cholesky constrains the numerical area of search and compromises the maximum likelihood theory currently used in behavioral genetics. Until the reasons for this phenomenon are understood and satisfactory solutions are developed, those who fit Cholesky matrices face the burden of demonstrating the validity of their fit statistics and the df for model comparisons. An interim remedy is proposed--fit an unconstrained model and a Cholesky model, and if the two differ, then report the difference in fit statistics and parameter estimates. Cholesky problems are a matter of degree, not of kind. Thus, some Cholesky solutions will differ trivially from the unconstrained solutions, and the importance of the problems must be assessed by how often the two lead to different substantive interpretation of the results. If followed, the proposed interim remedy will develop a body of empirical data to assess the extent to which Cholesky problems are important substantive issues versus statistical curiosities.

Algorithms↗
PubMed1986

The placental bed biopsy: review from three European centers.

This review derives from extensive experience with the placental bed biopsy technique in three centers over the last 30 years. A placental bed biopsy, usually taken at cesarean section, must include basal decidua and subjacent myometrium from the central zone of the placental site. Attention is drawn specifically to the sampling errors and to the pitfalls in morphologic interpretation of tissues, both maternal and fetal, that are continuously changing throughout the course of pregnancy. The features of the normal placental bed and of vascular lesions in pathologic pregnancies are briefly reviewed. Extension and elaboration of the technique and its more widespread use could contribute to the elucidation of many of the unresolved problems in human pregnancy.

Abortion, Spontaneous↗
PubMed1986

Resolution of steroid binding heterogeneity by Fourier-derived affinity spectrum analysis (FASA).

A new mathematical method of analyzing radioreceptor assay data is presented. When there are many binding classes with different affinities, the probability-density function B(p) is described by the equation B(p) = (integral negative infinity to infinity) q(k)f(p-k)dk, where q(k) is the affinity spectrum (density of a particular binding class as a function of affinity) and f(p-k) is a probability function (probability that dissociation constants will fall between k and p-k, where p is the free ligand concentration). This equation is solved for q(k) and evaluated explicitly by Fourier transformation, namely, q(w) = b(w)/f(w), where w is frequency. Since division by f(w) can amplify and high frequency noise present in the experimental data, a Gaussian smoothing function is introduced thus: qs(w) = q(w)e(-w/W0)2, where W0 is a constant. This produces an affinity spectrum defined as a plot of the number of binding sites, qs(k), versus their respective dissociation constants, k. Using a FORTRAN computer program, we verify this algorithm using simulated data. We also apply the procedure to resolve heterogeneous populations of estrogen binders in human endometrium using [3H]estradiol as ligand. Two estrogen binder classes are revealed with dissociation constants approximately 2.5 natural logarithmic units apart. We identify one high-affinity (Kd = 0.18 nM)-low density (70 pM [or 72 fmol/mg protein]) subpopulation and one low affinity (Kd = 2.5 nM)-high density (101 pM [or 102 fmol/mg protein]) subpopulation of estradiol binders. The management of experimental error, sampling limitations, and nonspecific binding are discussed. This method directly transforms experimental data into an easily interpretable representation without mathematical modeling or statistical procedures.

Endometrium↗
PubMed1992

Advanced computer programs for drug dosing that combine pharmacokinetic and symbolic modeling of patients.

In this paper, we describe our design for advanced drug dosing programs that "reason" using a combination of Bayesian pharmacokinetic modeling and symbolic modeling of patient status and drug response. Our design is similar to the design of the Digitalis Therapy Advisor program, but extends this previous work by incorporating a Bayesian pharmacokinetic model, performing a "meta-level" analysis of drug concentrations to identify sampling errors and changes in pharmacokinetics, and including the results of this analysis in reasoning for dosing and therapeutic monitoring recommendations. The design has been implemented in a program for aminoglycoside antibiotics called Aminoglycoside Therapy Manager. The program is user-friendly and runs on low-cost general-purpose hardware. The initial validation study showed that the program was as accurate in predicting future drug concentrations as an expert using commercial Bayesian forecasting software and that its dosing recommendations were similar to those of an expert.

Aminoglycosides↗
PubMed1980

Short latency somatosensory evoked potentials from radial, median and ulnar nerve stimulation in man.

Short latency somatosensory evoked potentials (SEPs) were elicited by stimulation at the wrist of median, radial, and ulnar nerves, singly or in combination, using normal subjects. Amplitude of P10 was strikingly lower with radial stimulation than with median stimulation, while ulnar-derived P10 was intermediate in amplitude. This difference probably reflects the antidromic firing of motor fibers contained in median nerves as compared with the superficial branch of radial nerve, which is entirely sensory. Beyond P10, there appear to be no significant differences between median, radial and ulnar-derived SEPs. With simultaneous stimulation of several nerves within one arm, larger potentials were sometimes achieved but with poorer definition of P12 and P14. The clinical utility of radial, ulnar, and median stimulation for localizing peripheral lesions derives from the distinct anatomical pathways of the stimulated fibers through the brachial plexus and from the separable motor and sensory components of P10. SEP is less invasive than EMG; this fact, plus its freedom from sampling error, make it potentially more suitable than conventional EMG for sequentially following a patient's clinical course.

Adult↗
PubMed1985

Histological measurement of fat content of liver of dairy cows.

A total of 275 liver biopsies were performed on dairy cows in 6 Friesian and one Guernsey herd during the first 2 weeks after calving. Liver samples were processed by 2 independent histological procedures: (a) formalin-fixed frozen sections were stained with oil-red O (ORO), or (b) samples fixed in glutaraldehyde-osmium tetroxide were embedded in plastic and sections stained with toluidine blue (TOLB). The sections were then subjected to stereological point-counting procedures to assess the quantities of stainable fat present within the liver cells. Estimates of liver cell fat by the 2 techniques were highly correlated, although those obtained by the TOLB method were consistently lower than those obtained by the ORO method. The analytical sampling error was slightly lower in the TOLB method. The simpler ORO method should prove an acceptable alternative in the routine histology laboratory to the tedious and technically demanding TOLB method.

Adipose Tissue↗
PubMed1985

Eye position during fixation tasks: comparison of macaque and human.

Two macaques and three humans fixated luminous targets in a dark field. All subjects had greater dispersion of eye position from trial-to-trial (between-trial variability) than would be predicted from sampling error and within-trial variability. Monkeys had greater between-trial dispersion on the vertical meridian than humans because of less precise control of saccades. Mean vertical eye position of the monkeys varied idiosyncratically with the fixation task (spot-dim or line-tilt). Between-trial fixation variability of both monkeys and humans was large enough to affect the interpretation of experiments relating visual performance to retinal anatomy or to neurophysiology.

Adult↗
PubMed1989

Immunoreactivity of tumor-associated glycoprotein (TAG-72) in normal, hyperplastic, and neoplastic colon.

Monoclonal antibody B72.3 recognizes tumor-associated glycoprotein (TAG-72) and has been widely used to identify malignant epithelial cells in cytologic and histologic preparations. We investigated TAG-72 expression in normal, hyperplastic, and neoplastic adult colon tissues. Formalin-fixed, paraffin-embedded samples of normal (43), hyperplastic (20), and neoplastic (70) colonic tissue were studied with B72.3 using the avidin-biotin complex immunoperoxidase method. TAG-72 expression was detected in 100% of the invasive carcinomas, in 84% of the normal colon samples, in 100% of the hyperplastic polyps, and in 93% of the adenomatous or mixed hyperplastic-adenomatous lesions. Among cases expressing TAG-72 immunoreactivity, the extent of staining varied from 100% of cells in normal and hyperplastic lesions to 10% to 100% in neoplastic lesions. A consistent supranuclear (presumably Golgi) staining pattern was observed in all tissues with the exception of carcinoma and some adenomas; in these cases, staining localized to the luminal surface and intraluminal mucin and/or was diffusely cytoplasmic. From these data, it is clear that TAG-72 is expressed in both benign and neoplastic cells of the human adult colon, although different patterns of expression may distinguish malignant transformation. Furthermore, carcinoma cells less consistently express TAG-72, so that small biopsies or cytologic specimens may be falsely negative due to sampling errors.

Adenoma↗
PubMed1992

Radiation therapy for nasopharyngeal carcinoma: histologic appearances and patterns of tumor regression.

Nasopharyngeal carcinoma is a common malignancy in Hong Kong and is treated by external radiotherapy. After 6.5 weeks of radiotherapy, the nasopharynx of 100 patients was examined and biopsy specimens were taken. All patients had repeated examination and biopsies done every 2 weeks until exophytic tumor was not seen and biopsy samples were negative on more than one examination of the nasopharynx. The interval between the cessation of therapy and biopsy ranged from 1 day to 11 weeks. Twenty-three patients had atypical findings and five of these had residual tumor requiring gold grain implantation brachytherapy. We identified a number of distinct pathologic changes in the post-biopsy material. Most of these changes disappeared 8 weeks after the cessation of therapy, but the presence of residual tumor after this time was an indication for subsequent therapy. If the date of the first biopsy was delayed until 6 weeks after the completion of radiotherapy, the percentage of atypical biopsies containing residual tumor rose from 21% to 55%. The posttreatment biopsy should be performed twice, as four patients had negative biopsy findings due to sampling error.

Adolescent↗
PubMed1989

Prospective analysis of liver biopsies before and after methotrexate therapy in rheumatoid patients.

The significance of hepatic changes in methotrexate-treated RA patients is unclear at this time. In our group of RA patients, there was a slight increase in the incidence of triaditis and fat during methotrexate therapy. Disease duration greater than or equal to 10 years was associated with increased hepatic triaditis before treatment. Age greater than 50 years was associated with increased hepatic fat before and after treatment. It appears that patients' ages and duration of underlying RA account for some changes, independent of methotrexate therapy. Several of our patients changed from higher to lower histologic grade or had an apparent decrease in fibrosis, fat, or triaditis on the pathologists' reports and the blind readings of the repeat biopsies. This may be explained by sampling error. More importantly, some of these changes may not be of clinical significance. One report of methotrexate-induced cirrhosis in patients with psoriasis demonstrated that in all but one of 14 patients who continued receiving methotrexate the cirrhosis decrease or did not progress. This may also be true of the hepatic fibrosis seen in RA after methotrexate treatment. In this study, there did not appear to be changes seen on pretreatment liver biopsy that were predictive of subsequent fibrosis or cirrhosis. Our data indicate that pretreatment biopsy is unwarranted in a population similar to ours. However, our practice has been to try to avoid methotrexate in patients with diabetes, prior liver disease, alcoholism, or obesity because of previous reports suggesting that these patients are at increased risk for the development of cirrhosis. Only the above-mentioned patient, eventually diagnosed as having cirrhosis, might have been handled differently. Including the study, none of the approximately 700 RA patients in the literature having liver biopsies after methotrexate therapy have developed cirrhosis consequent to its use. Most of these had received a total dose of approximately 1,500 mg in small weekly doses, and alcohol was prohibited. Below this cumulative dose the risk of clinically significant liver damage in carefully selected patients is very low. In view of this experience, the recommendation that RA patients have liver biopsies after 1,500 mg of methotrexate (a holdover from the psoriasis literature) may be too conservative in low-risk RA patients, provided methotrexate is administered weekly and alcohol is prohibited. Recognizing that the absolute need for biopsy is unproven, a more realistic milestone for those choosing biopsy might be after each 2,000 to 2,500 mg.(ABSTRACT TRUNCATED AT 400 WORDS)

Aged↗
PubMed1991

The accuracy of frozen section diagnosis of ovarian tumors.

We compared all frozen section examinations of ovarian tumors during a 6-year period in our institute with the final diagnosis from paraffin sections. In this period, 946 ovarian tumor specimens were removed for histologic assessment; 176 (18.6%) had frozen section examination. Final histological diagnosis was divided into benign (55.1%), borderline malignant (10.3%), and malignant (34.6%). Sensitivity of the frozen section method for malignant or borderline disease was 83.5% and specificity for a benign lesion, 92.8%. Predictive values and 95% confidence intervals were computed: 100% (93-100%) for malignancy, 62% (32-86%) for borderline malignancy, and 92% (85-96%) for a benign disease. Diagnostic problems occurred in large borderline tumors of mucinous cell type. Analysis of the 12 false negative diagnoses revealed that a sampling error was involved in 11 cases. A judgment error was made in the only false positive and in 1 out of 12 false negative frozen section diagnoses. It is concluded that when surgeons and pathologists are aware of the limitations of frozen section diagnosis of ovarian tumors, peroperative histologic examination can be worthwhile and prevent under- and overtreatment of gynecologic patients.

Adolescent↗
PubMed1993

Proposal for a classification of acute myeloid leukaemia based on plastic-embedded bone marrow biopsy sections.

Traditional approaches to haematological diagnosis of acute myeloid leukaemia (AML) include microscopic examination of peripheral blood (PB) and bone marrow (BM) cells studied in Romanowsky-stained, dry film smears. The routine evaluation of BM aspirates, however, may be disappointing in accurately assessing marrow activity because of the sampling error inherent in the BM aspiration technique, and occasionally may fail to provide a precise diagnosis because BM aspirates cannot be obtained due to packed or fibrotic marrows. Because of the advantages it offers over the conventional method of diagnosis, we have studied Romanowsky-stained, thin sections of plastic-embedded BM biopsies from 87 newly diagnosed AML patients. Based on our study, the patients were broadly grouped into four categories: (1) those with hypocellular marrow (haemopoietic cellularity less than 50%), hypoplastic AML; (2) those with some degree of marrow fibrosis, AML with marrow fibrosis; (3) those with homogeneous infiltration of marrow with blast cells; and (4) those with inhomogenous infiltration of marrow with blast cells. Our results show that the proposed classification of AML based on the evaluation of plastic-embedded BM biopsy sections may offer benefits over those afforded by conventional morphologic smear techniques. This may provide significant prognostic information and may have a major impact on patient management. It may also provide a meaningful stratification of the patient population when comparing treatment protocols for newly diagnosed AML patients.

Biopsy, Needle↗
PubMed1980

The use of immunochemically purified anti-brucella antibody in a direct fluorescent antibody test for Brucella abortus.

Antibodies specific for Brucella abortus were purified from the serum of hyperimmunized sheep using immunochemical procedures. They were conjugated to fluorescein isothiocyanate and used in a fluorescent antibody (FA) test for B. abortus. The conjugate did not stain any heterologous bacterial or fungal species tested and background fluorescence associated with its use on smears and sections of abortion materials was particularly low. Of 239 cases of abortion examined fluorescent microscopy demonstrated B. abortus in all 12 cases in which the organism was isolated. A few areas of fluorescence typical of B. abortus were also seen in 3 cases from which the organism was not cultured. B. abortus was demonstrated in lymph nodes from 6 of 36 Brucella reactor cows by culture and 7 by the FA test. However, only very low numbers of B. abortus were isolated or seen and sampling errors would have been significant. Use of the FA test allows diagnosis of Brucellosis to be made in 2 hours, compared to 6 days by the usual cultural procedures.

Aborted Fetus↗
PubMedDate not supplied

Filter in situ hybridisation: an evaluation of the FISH technique for HPV detection in cervical swabs.

The filter in situ hybridisation (FISH) method for detection of HPV in cervical swabs was evaluated against the Southern blot technique on concomitant cervical biopsies. Of 73 biopsies, HPV 16 DNA sequences were found in 26 biopsies and HPV 18 sequences in 2 biopsies. Analysis by FISH of the corresponding smears detected 58 and 100% of these, respectively. Of the smears corresponding to the HPV-negative biopsies, 17% were HPV 16-positive and 3% were HPV-18 positive by FISH. Re-hybridisation with cold plasmid added for competition did not change these results. To estimate the risk of spurious hybridisation between vector remnants in the probe and bacterial DNA sequences present in smears, we have hybridised by FISH to preparations of the 19 most common vaginal microorganisms. Of these, E. coli, which is present in about 10% of cervical smears, hybridised strongly with a probe of the plasmid vector pBR322 and may be a significant cause of false positive FISH results. None of the bacteria hybridised with probes of purified HPV when cold, denatured plasmid was added for competition. Analysis by FISH with probes of purified pBR322 to 167 smears of a patient control group resulted in 6% positive reactions. In hybridisations with probes of HPV 16 and 18 to 2 or 3 different filter preparations of the same smear, identical results were obtained in 18 of 19 smears, indicating a good reproducibility by the FISH method. The high percentage of HPV negative smears is equivalent to the rates known from cytology and may reflect sampling errors.

Biopsy↗
PubMed1995

Healthy human T-cell receptor beta-chain repertoire. Quantitative analysis and evidence for J beta-related effects on CDR3 structure and diversity.

Analysis of TCR repertoire usage and clonality is of potential value in understanding the pathogenesis of a number of human immune-mediated diseases. In diseases that are likely to be dependent on antigen-driven T cells, it has been suggested that particular TCR junctional region or CDR3 sequences may be critical. Rigorous methods for TCR analysis which are both quantitative and qualitative are therefore required. Of those commonly available, only anchor and inverse PCR are capable of giving high-quality information on V, D, N, and J region usage, but it has not been established whether both methods are quantitatively or analytically equivalent. We show here that both methods detected considerable variability in the usage of V beta and J beta segments in the peripheral blood repertoire of a normal individual. No preferential V-J pairing could be demonstrated. An excess usage of J beta 2 family members was indicated by both methods, although the relative usage of different J beta 2 families differed between the two techniques. The predominantly used V beta usage showed that for some families, estimates of their frequency in the repertoire differed significantly between the anchor and inverse libraries. When sampling relatively few clones the variation between V beta families estimated using the two methods can be considerable. This is likely to be a result of sampling error from a large gene family. Large-scale screening of several thousand clones is recommended to confirm the absolute values obtained from sequencing. Variation in CDR3 length appeared to be normally distributed, suggesting that a statistically optimal junctional region length may have been selected for contact with antigen. CDR3 length distribution differs significantly between receptors, which have rearranged to J beta 1 versus J beta 2 families, with the J beta 2-associated CDR3 on average between 0.5 and 1.2 of an amino acid longer. Thus the TCR beta junctional region repertoire of humans is subject to structural constraints associated with J beta usage. It will be important to establish whether variation in CDR3 length and J beta usage exists between subsets of human T cells in order to interpret TCR repertoire data from disease and control tissues.

Adult↗
PubMed1988

The role of aspiration cytology in the management of thyroid nodules.

In the hands of an experienced cytologist aspiration cytology is a safe and hitherto the best diagnostic tool in the evaluation of nodular thyroid lesions. In histologically verified case series 50-90% of confirmed thyroid cancers can be detected by aspiration biopsy, the sensitivity being dependent on sampling errors, microscopic misinterpretation and the variation in attitude towards indeterminate diagnosis in the decision for diagnostic surgery. The number of proven benign cases that are correctly identified as such by biopsy varies accordingly and approx. 75% (specificity). In comparison with imaging procedures, including those giving information of functional activity, the combined sensitivity and specificity rates of aspiration cytology come closest to the ideal discriminatory situation. In combination with case history and careful clinical examination, fine needle aspiration cytology is the best guidance for an optimal selection of patients for therapeutic or diagnostic surgery. Future development of sensitive markers for malignant degeneration will probably increase the selective power of this diagnostic technique.

Biopsy, Needle↗