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Serum standards for the bioassay of aminoglycosides in cerebrospinal fluid.

Four diluents were compared as reference standards for the assay of gentamicin in cerebrospinal fluid (CSF): human CSF, human serum, distilled water, and 150 mmol NaCl/4.5 mmol CaCl2. Standards prepared in pooled human serum were the best alternative to CSF for the assay of gentamicin and were also useful for the assay of tobramycin, netilmicin, amikacin, and sisomicin. The pH (6.0-9.8) of CSF did not alter the results of the assay.

Aminoglycosides

Which radiographic plane should be used to quantify the distal tibia angle on weightbearing CT images?

BACKGROUND: Precise quantification of distal tibial alignment is essential for planning corrective osteotomies and ankle joint replacement surgery. The lateral distal tibial angle (LDTA) is the principal radiographic parameter used for this purpose. While LDTA is increasingly measured on weightbearing cone-beam CT (WBCT) using two-dimensional coronal slices, the optimal measurement plane remains unclear. METHODS: In this retrospective comparative study, full-leg WBCT scans of patients scheduled for supramalleolar osteotomy (n&#x202f;=&#x202f;20; mean age 47&#x202f;&#xb1;&#x202f;12.8 years) were analyzed. LDTA was measured on three coronal planes of the distal tibial plafond (anterior edge, mid-dome, posterior edge) and compared with semi-automated three-dimensional (3D) tibial alignment measurements as the reference standard. RESULTS: Mid-dome LDTA showed no significant difference from the 3D reference (p&#x202f;>&#x202f;0.05) and demonstrated excellent agreement. Anterior measurements significantly overestimated LDTA, while posterior measurements underestimated it (both p&#x202f;<&#x202f;0.05), with only fair agreement. CONCLUSION: LDTA should be measured at the mid-dome of the distal tibial plafond on WBCT to ensure accurate and reproducible alignment assessment. LEVEL OF EVIDENCE: Level III - Retrospective Comparative Study.

Humans

Comparison of protein values in cerebrospinal fluid by nephelometry and radial immunodiffusion.

Various parameters of three radial immunodiffusion systems, a kinetic nephelometer and two laser nephelometers were compared in a measurement of IgG and albumin in cerebrospinal fluid. Greater precision for IgG and albumin determinations was found with two RID systems. Correlation coefficients between methods ranged from 0.71 to 0.94. The generally poor correlations of methods, combined with the results of accuracy studies employing WHO standard reference dilutions and proportional errors demonstrated by linear regression equations, illustrate graphically the need for standardization in the field of immunochemical determinations. Our preliminary studies of the IgG/Alb ratio in cerebrospinal fluid of multiple sclerosis patients agreed with the work of others. In each case, the ratio was elevated or borderline high.

Albumins

Application of a spectrophotometric method to the determination of the composition of oligonucleotides obtained from cysteine transfer ribonucleic acid.

1. The applications of methods for determining the composition of oligonucleotides from u.v.-absorption spectra is described. 2. In the first method absorbances at selected wave-lengths were read from the spectra of oligonucleotides in solution in 7 M-urea which had been recorded at acid and alkaline pH values. 3. In the second method absorbances were sampled automatically at regular time-intervals during scans at acid and alkaline pH of each spectrum, converted into digital signals and recorded on paper take for computer processing. The holmium spectrum in the region of the holmium peak at 333.7 nm was superimposed on each nucleotide spectrum. The position of this peak maximum was used as a standard reference point in the computer-based analysis. 4. By using either method the composition was calculated by a least-squares procedure by using a library of values for five standard nucelotides obtained in a similar manner. 5. The methods gave satisfactory compositions for mixtures of mononucleotides as well as for five dinucleoside monophosphates. 6. Methods of minimizing the effects on the nucleotide composition of spectural changes due to base stacking are discussed. 7. The compositions of some oligonucleotides obtained during an investigation of the nucleotide sequence of tRNA (Cys) were determined and agreed with the sequences found by other methods.

Base Sequence

Use of indocyanine green fluorescence versus patent blue V dye for sentinel lymph node biopsy in early breast cancer, a randomized controlled trial.

BACKGROUND: Sentinel lymph node biopsy (SLNB) is standard for axillary staging in early breast cancer. While the combination of radioisotope and blue dye (e.g., patent blue V, PBV) remains the standard, it has limitations including logistics, variable identification rate (IR), and allergic potential. Indocyanine green (ICG) fluorescence is a promising alternative, but high-quality comparative evidence is needed. METHODS: This was a single-center, prospective, randomized controlled trial. Forty patients with early-stage, node-negative breast cancer were allocated to SLNB using either ICG (n&#x2009;=&#x2009;20) or PBV (n&#x2009;=&#x2009;20). All patients subsequently underwent completion level I-II axillary lymph node dissection (ALND) as the pathological reference standard for diagnostic performance assessment. Primary outcome was sentinel lymph node (SLN) IR. Secondary outcomes included detection time, number of SLNs retrieved, false-negative rate (FNR), and safety. RESULTS: Baseline characteristics were comparable between groups. The SLN IR was significantly higher with ICG (100% [20/20]) than with PBV (75% [15/20], p&#x2009;=&#x2009;0.047). ICG was associated with a significantly shorter median detection time (14.5 vs. 24.0&#xa0;min, p&#x2009;<&#x2009;0.001) and retrieved more SLNs (mean: 3.6 vs. 2.4, p&#x2009;=&#x2009;0.002). Most critically, ICG demonstrated 100% sensitivity, specificity, negative predictive value (NPV), and overall diagnostic accuracy, with a 0% FNR. In contrast, PBV achieved a sensitivity of 75%, an overall diagnostic accuracy of 90%, and an FNR of 25%. No ICG-related adverse events occurred. PBV caused skin discoloration in 75% of patients and one (5%) allergic reaction. CONCLUSION: ICG fluorescence achieved a higher SLN IR, shorter detection time, higher sensitivity, lower FNR, and fewer tracer-related adverse events than PBV as a single tracer for SLNB in patients with early-stage breast cancer. These findings suggest that ICG is a promising standalone tracer when radioisotope mapping is unavailable. Larger multicenter studies are required before widespread adoption can be recommended.

Humans

Comparison studies on potency tests for rabies vaccines.

Eleven laboratories from eight countries and four continents took part in a collaborative study to evaluate experimental procedures to be used in selecting the new standard reference rabies vaccine prepared in human diploid cell cultures. The following procedures were used : (a) the NIH potency test in mice, (b) the antibody binding technique (by either mouse inoculation or the tissue culture method), (c) virus-neutralizing antibody levels in mice used for the NIH test, and (d) antibody induction in human volunteers treated with vaccine alone and in combination with human rabies gamma globulin. The four methods used for determination of rabies antibodies were mouse inoculation, rapid fluorescent focus inhibition, plaque reduction and complement fixation techniques. All results were expressed in International Units as compared to the standard WHO serum and vaccine preparations. In general, a close correlation was noted in results from different laboratories, and it was recommended that the future rabies standard vaccine should be evaluated by three testing procedures, the NIH test, the antibody-binding technique, and antibody levels in mice used for the NIH test.

Animals

Characteristics of p53 and Smad4 immunohistochemistry in pancreatic ductal adenocarcinoma and validation by next-generation sequencing.

BACKGROUND: Mutations in four major driver genes -KRAS, CDKN2A, TP53, and SMAD4- are central to the pathogenesis of pancreatic ductal adenocarcinoma (PDAC) and critically inform diagnosis, therapeutic decision-making, and prognostic assessment. Although next-generation sequencing (NGS) is widely regarded as the gold standard for detecting these mutations, its clinical application is often limited by suboptimal analytical efficiency and substantial economic cost. Among these genes, immunohistochemical (IHC) staining for the proteins encoded by TP53 and SMAD4 has been extensively adopted in routine pathology practice. However, standardized IHC pattern classification schemes and rigorous validation of their predictive accuracy for underlying genomic alterations remain lacking in PDAC. METHODS: We retrospectively enrolled 63 PDAC patients and systematically characterized the typical IHC expression patterns of p53 and Smad4. Targeted NGS was subsequently performed on all available tumor specimens, and the resulting mutational profiles were correlated with corresponding IHC findings. Diagnostic performance including sensitivity, specificity and accuracy of p53 IHC for predicting TP53 mutations and of Smad4 IHC for predicting SMAD4 mutations was rigorously evaluated. RESULTS: Among the four canonical driver genes, co-occurring double- or triple-gene mutations were prevalent; within TP53 and SMAD4, missense mutations constituted the most frequent variant type. Using NGS as the reference standard, we validated the diagnostic utility of a three-tiered p53 IHC classification system, particularly in fine-needle biopsy (FNB) specimens. Furthermore, we proposed a novel, refined Smad4 IHC pattern classification that incorporates an "intermediate" category, thereby expanding upon conventional binary interpretation. This new scheme achieved markedly improved mutation prediction accuracy (0.76) compared with traditional approaches (0.57). CONCLUSION: Our study highlights the complementary diagnostic value of p53 and Smad4 IHC relative to molecular testing in PDAC, especially when tissue is limited, as commonly encountered in FNB specimens. The newly established Smad4 IHC classification system, which integrates an intermediate expression category into the conventional two-tier framework, demonstrates superior clinical utility and enhances predictive accuracy for SMAD4 genomic alterations.

Humans

Observer variation in the interpretation of lower limb venograms.

After agreeing on diagnostic criteria and after a pilot study, two experienced radiologists twice independently reviewed 40 lower limb venograms performed by a standard technique in patients suspected or known to have venous thrombosis. The observers reviewed 20 examinations at a time, their analysis requiring separate identification of 11 major veins. At each site observers stated whether thrombus was "absent," "doubtful," "presumed," or "definite," or declared "no opinion possible." They then rediscussed criteria of diagnosis and, using the same experimental design, examined another 40 venograms. To correct for agreement expected by chance, data were analyzed by using the kappa statistic. In general, levels of agreement were higher than those reported for many other clinical and radiologic investigations, probably because of refinement of criteria after the pilot study. Nonetheless, observers disagreed about the probable presence or absence of thrombus at some site in the limb in about 10% of examinations. Observer variation should be considered when venography is used as a reference standard to evaluate other methods of diagnosing thrombi.

Adult

Stretched penile length in boys with hypospadias: Population-based analysis using validated nomogram.

BACKGROUND: Hypospadias affects 1 in 200-300 male births. Parents are often concerned about penile adequacy beyond the urethral defect itself, yet few studies have systematically compared stretched penile length (SPL) in hypospadias against population-based reference standards. OBJECTIVE: To evaluate SPL distribution patterns in boys with Types I and II hypospadias and compare them with established normative data. METHODS: The authors studied 876 consecutive boys aged 1-14 years with unoperated Types I (distal) and II (mid-shaft) hypospadias. Two observers independently measured SPL using the validated SPLINT technique. The SPL measurements were compared against age-matched normative data from 1276 Indian children. Exact binomial probability tests were used for percentile distributions, chi-square tests for subtype comparisons and t-tests for mean deviations. RESULTS: The cohort included 479 Type I and 397 Type II cases. SPL distribution showed a marked leftward shift: 71% fell below the 50th percentile (expected 50%, p < 0.001) and 41.5% below the 25th percentile. Lower percentiles were overrepresented, 20.7% were below the 10th percentile and 20.8% in the 10th-25th range. Upper percentiles were depleted: only 7.4% in the 75th-90th range and 1.7% above the 90th percentile (all p < 0.001). Mean SPL was reduced by 6.8% (95% CI: -8.18 to -5.42%) in Type I and 7.5% (95% CI: -9.05 to -5.92%) in Type II. The two subtypes showed no significant distributional difference (&#x3c7;2 = 6.22, p = 0.18), suggesting that meatal position does not predict SPL reduction. CONCLUSIONS: Boys with distal and mid-shaft hypospadias show clinically meaningful SPL reduction that follows a continuous distribution rather than an all-or-none pattern. SPL reduction appears independent of meatal position. These findings support routine SPL assessment using population-specific references and can guide preoperative counselling.

Humans

Soft Tissue Volume Augmentation at Single Implant Sites Applying Collagen Matrices or Connective Tissue Grafts: 10-Year Follow-Up of a Randomized Controlled Trial.

AIM: To compare up to 10&#x2009;years clinical, profilometric and patient-reported outcomes of implant sites previously augmented using a volume-stable collagen matrix (VCMX) or connective tissue graft (SCTG) in the aesthetic zone. METHODS: The original non-inferiority randomized controlled trial (RCT) enrolled 20 patients who received soft tissue volume augmentation with VCMX or SCTG at single implant sites. Clinical assessments and standardized measurements were performed at baseline after crown insertion and at 6&#x2009;months, 1, 3, 5, 7.5, and 10&#x2009;years. The primary outcome was mucosal thickness. Secondary outcomes included marginal bone levels (MBL), probing depth (PD), bleeding on probing (BOP), plaque control record, Pink Aesthetic Score (PES), OHIP-14 and buccal profilometric changes. Group comparisons were performed using mixed-effects and generalized estimating equation (GEE) models, which account for within-patient correlations due to repeated measurements and allow inclusion of all available data without requiring imputation for missing observations. RESULTS: Of the 20 originally enrolled patients, 10 (5 in the SCTG group and 5 in the VCMX group) were available for re-examination at 10&#x2009;years. The adjusted between-group difference in mucosal thickness was -0.02&#x2009;mm (95% CI -0.99 to 0.96). As the lower bound of the confidence interval remained above the prespecified non-inferiority margin of -1&#x2009;mm, non-inferiority of VCMX was shown. Buccal contour changes were comparable during the early follow-up, while a trend toward a greater long-term contour decrease was observed in group VCMX (-0.31&#x2009;mm [95% CI, -0.65 to 0.03]; p&#x2009;=&#x2009;0.07). Mean PES values were 10.6 in the SCTG group and 9.6 in the VCMX group, with no significant between-group differences (p&#x2009;=&#x2009;0.45). Both groups revealed high levels of oral health-related quality of life, with low median OHIP-14 scores (SCTG, 0.0; VCMX, 1.0; p&#x2009;=&#x2009;0.26). CONCLUSION: These preliminary long-term findings showed no clinically relevant differences between SCTG and VCMX in terms of clinical, profilometric and patient-reported outcomes. While SCTG remains the reference standard, VCMX represents a less invasive alternative but with a slight tendency toward greater long-term contour reduction. CLINICAL SIGNIFICANCE: Volume-stable collagen matrices serve as a viable alternative to autogenous connective tissue grafts for peri-implant soft tissue volume augmentation, particularly in patients seeking a reduced morbidity, without compromising long-term clinical or aesthetic outcomes. TRIAL REGISTRATION: German Clinical Trials Register: DRKS00017484.

Humans

Home screening for the detection of urinary tract infection in infancy.

One hundred sixty-five infants of both sexes, 2 weeks to 2 years of age, were screened by their parents at home for bacteriuria. Parents were instructed during a single brief period during the initial office visit to collect and to culture their infants' urine at home. Urine was collected by specimen bag and cultured promptly at cribside using a newly developed culture kit and inoculating swab. The same specimen was cultured by the office laboratory staff by kit and swab and, as a reference standard, by the loop-MacConkey method. Infants were screened at home after three consecutive office visits. Then, 870 specimens were processed. Urinary tract infection (UTI) was diagnosed in 3.6% of the study population. Results show that home screening of infants for UTI by the parent compares favorably with screening by the office laboratory.

Bacteriological Techniques

Diagnostic Accuracy of a CRISPR-Based Assay in Smear- and Culture-Negative Fungal Keratitis.

IMPORTANCE: Diagnosing fungal keratitis (FK) in patients with negative smear and culture results remains clinically challenging, highlighting the need for alternative diagnostic approaches. OBJECTIVE: To determine the diagnostic accuracy of the clustered regularly interspaced short palindromic repeats (CRISPR)-based Rapid Identification of Mycoses using CRISPR (RID-MyC) assay for detecting FK in patients with negative smear and culture results using in vivo confocal microscopy (IVCM) as the reference standard. DESIGN, SETTING, AND PARTICIPANTS: This prospective diagnostic accuracy study was conducted from December 2024 to March 2025 at Aravind Eye Hospital, a tertiary ophthalmology referral hospital in Coimbatore, India. Consecutive patients clinically suspected to have microbial keratitis with negative smear and culture results were eligible for inclusion. Data were analyzed from March 2025 to June 2025. INTERVENTIONS: All included participants underwent corneal scraping for RID-MyC assay and imaging by IVCM. MAIN OUTCOMES AND MEASURES: The primary outcomes were sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic concordance of the RID-MyC assay compared with IVCM results. RESULTS: Of 245 consecutive patients clinically suspected to have microbial keratitis, 82 were smear negative. After exclusions due to contraindications or positive subsequent cultures, 41 patients with smear- and culture-negative results were ultimately included in the final analysis. Of these 41 patients (mean [SD] age, 51.0 [14.6] years; 21 [51.2%] women), RID-MyC demonstrated sensitivity of 82.1% (95% CI, 63%-94%) and specificity of 76.9% (95% CI, 46%-95%). Positive predictive value was 88.5% (95% CI, 74%-95%) and negative predictive value was 66.7% (95% CI, 46%-82%). Concordance between RID-MyC and IVCM was observed in 33 cases (80.5%). Notably, prior antifungal treatment was most frequent (4 of 5 [80%]) among patients with positive IVCM but negative RID-MyC results. Conversely, all patients (3 of 3 [100%]) with negative IVCM but positive RID-MyC findings had smaller, peripheral, or paracentral lesions. CONCLUSIONS AND RELEVANCE: In this diagnostic study, in patients with smear- and culture-negative FK, the RID-MyC assay showed good diagnostic accuracy comparable with IVCM and was feasible in all cases, including those in whom imaging was not possible. With its rapid turnaround and minimal equipment needs, RID-MyC may serve as a practical adjunct to conventional diagnostics, particularly in high-burden, resource-limited settings where IVCM is unavailable or contraindicated.

Humans

Holmium laser enucleation of the prostate for the treatment of lower urinary tract symptoms in men with benign prostatic hyperplasia.

RATIONALE: A range of surgical options is available for the treatment of benign prostatic hyperplasia (BPH), including holmium laser enucleation of the prostate (HoLEP). The evidence is unclear regarding differences in functional, perioperative, and morbidity outcomes between these modalities. OBJECTIVES: To assess the effects of holmium laser enucleation of the prostate compared with other surgical treatments for lower urinary tract symptoms in men with benign prostatic hyperplasia. SEARCH METHODS: We searched multiple databases (including MEDLINE, Embase, CENTRAL, Web of Science, LILACS, and the International HTA database), trial registries, and conference abstracts through April 08, 2026. ELIGIBILITY CRITERIA: We only included randomized trials of men over 40 years of age with a prostate volume of at least 20 mL (assessed by digital rectal examination, ultrasound, or conventional imaging) who exhibited lower urinary tract symptoms (LUTS) defined by an International Prostate Symptom Score (IPSS) of eight or greater undergoing surgical interventions for BPH. OUTCOMES: The critical outcomes measured were the urologic symptoms score, the quality-of-life score, and major adverse events. The important outcomes measured were: re-treatment, erectile function, ejaculatory function, transfusions, acute urinary retention, indwelling urinary catheter duration, and hospital stay duration. RISK OF BIAS: We used the Cochrane risk of bias tool (RoB 1) to assess for potential sources of bias on a study and outcome level basis. SYNTHESIS METHODS: We pooled outcome data using the random-effects model and performed meta-analyses using the Mantel-Haenszel method. We assessed statistical heterogeneity in the pooled data by visually inspecting forest plots and using the I2 statistic to quantify it. We used the GRADE framework to assess the certainty of evidence. INCLUDED STUDIES: We included 52 trials that included 6242 participants that compared HoLEP to other surgical interventions for benign prostatic hyperplasia. The median age of participants across the studies ranged from 65 to 74 years. The baseline prostate volume ranged from 30 cc to 142 cc. Baseline IPSS scores ranged from 19.6 to 28.6 (range 0-35). SYNTHESIS OF RESULTS: We prioritized comparing HoLEP with transurethral resection of the prostate (TURP) at short-term follow-up (up to 12 months), because TURP is the long-standing reference standard and the predominant comparator in randomized surgical trials. Findings for the four remaining comparisons (laser ablation, alternative energy source enucleation, other minimally invasive therapies, and simple prostatectomy), for long-term follow-up, and for all remaining outcomes are reported in full in the review. Compared to TURP, at short-term follow-up: Critical outcomes - HoLEP may result in little to no difference in short-term urologic symptom scores measured using the IPSS (range 0 to 35; lower values reflect fewer symptoms) (MD -0.67, 95% CI -1.20 to -0.14; I&#xb2; = 93%; 14 studies, 1666 participants, low-certainty evidence). - HoLEP may result in little to no difference in short-term quality of life (range 0 to 6; lower values reflect better quality of life) (MD -0.04, 95% CI -0.23 to 0.15; I&#xb2; = 73%; 6 studies, 876 participants, low-certainty evidence). - HoLEP may result in little to no difference in short-term major adverse events (RR 0.75, 95% CI 0.35 to 1.58; I&#xb2; = 0%; 10 studies, 1147 participants, low-certainty evidence). Important outcomes - HoLEP likely results in little to no difference in re-treatment (RR 0.45, 95% CI 0.14 to 1.50; I&#xb2; = 0%; 8 studies, 813 participants, moderate-certainty evidence). - HoLEP likely results in little to no difference in erectile function (MD -0.03, 95% CI -0.47 to 0.42; I&#xb2; = 0%; 3 studies, 518 participants, moderate-certainty evidence). - Ejaculatory function: we did not find any data for this outcome. - HoLEP likely reduces the need for blood transfusion (RR 0.19, 95% CI 0.09 to 0.42; I&#xb2; = 0%; 15 studies, 1755 participants, moderate-certainty evidence). AUTHORS' CONCLUSIONS: Compared with TURP, HoLEP may achieve similar relief of urologic symptoms, similar quality of life, and similar rates of major adverse events in the first 12 months after surgery, and probably similar re-treatment rates and erectile function. HoLEP likely reduces the need for blood transfusion; this is the only advantage of HoLEP that the randomized evidence, as summarized here, supports as clinically important. There was insufficient evidence to assess outcomes in the subset of individuals with larger prostates or on anticoagulation. Future research should prioritize long-term trials reporting sexual function and urinary incontinence outcomes, recruit men with very large prostates (&#x2265; 150 cc) or on anticoagulation therapy, and evaluate cost-effectiveness and training requirements. FUNDING: No external funding was received for this review. REGISTRATION: The protocol for this review was published in the Cochrane Database 2019 (https://doi.org/10.1002/14651858.CD013291).

Humans

Size and pericentric inversion heteromorphisms of secondary constriction regions (h) of chromosomes 1, 9, and 16 as detected by CBG technique in Caucasians: classification, frequencies, and incidence.

Eighty normal Caucasians were studied by CBG technique for estimation of size and inversion heteromorphisms of chromosomes 1, 9, and 16. Size heteromorphisms were classified into one of five sizes using 16p as a reference standard: very small, small, intermediate, large, and very large. Inversion heteromorphisms were also classified into 5 categories - eg, no inversion; partial inversion - minor; half inversion; partial inversion - major; and complete inversion. The frequencies of size heteromorphisms for chromosomes 1, 9, and 16 were 11.3%, 47.5%, and 7.5%, respectively. Thirty-four chromosomes were found to have inversions. Of these, 16 were in chromosome 1, and 18 were in chromosome 9. No inversions were found in chromosome 16. An increase in the size of the h region was more frequently associated with inversion, suggesting that there is a possible relationship between size and inversion. For example, there were 118 chromosomes that were classified as "intermediate" by size; 23 (19.5%) had inversions. In contrast, there were 225 that were "small" in size, and only 10 (4.4%) had inversions. There was no significant difference between males and females for size and position heteromorphisms.

Adult

Absolute Quantification of Cellular and Cell-Free Mitochondrial DNA Copy Number from Human Blood and Urinary Samples Using Real Time Quantitative PCR.

Mitochondrial DNA copy number (mtDNA-CN) in human body fluids is widely used as a biomarker of mitochondrial dysfunction in common metabolic diseases. Here we describe protocols to measure cellular and/or cell free (cf)-mtDNA-CN in human peripheral blood and urine. Cellular mtDNA is located inside the mitochondria where it encodes key subunits of the respiratory complexes in mitochondria and is usually normalized with reference to the nuclear genome as the mitochondrial genome to nuclear genome ratio (Mt/N) in either whole blood, peripheral blood mononuclear cells (PBMCs), or whole urine. Cf -mtDNA is usually found outside of the mitochondria, often released following mitochondrial damage, can trigger inflammatory pathways, and is usually measured as mtDNA-CN per volume of the starting material. Here we describe how to (1) separate whole blood into PBMCs, plasma, and serum fractions and whole urine into urinary supernatant and pellet, (2) prepare DNA from each of these fractions, (3) prepare reference&#xa0;standards&#xa0;for absolute quantification, (4) carry out qPCR for either relative or absolute quantification from test samples, (5) analyze qPCR data, and (6) calculate the sample size to adequately power studies. The protocol presented here is suitable for high throughput use and can be modified to quantify mtDNA from other body fluids, human cells, and tissues.

Humans

[Isolation and characterization of pregnancy associated alpha2 glycoprotein].

A method using immunoadsorbents for the isolation of pregnancy-associated alpha2-glycoprotein (alpha2-PAG) from the extract of human placentae is described. The physical properties and the chemical composition of the purified protein are determined: alpha2PAG sediments with 11,5 S, has a molecular weight of 360 000 daltons and is composed of subunits having a molecular weight of 180000, which are held together by disulfide bonds. The isoelectric point was found to be pH 4,7 and the extinction coefficient (E1%1cm) was determined to be 9,7 at 277 nm. The carbohydrate content of the molecule amounts to 12,1% (hexose 6,0%, hexosamine 3,7%, fucose 0,06%, sialic acid 2,4%). An analysis of the amino acids is reported, too. The purified alpha2PAG was used to determine the absolute concentrations of this protein in a reference standard and in sera.

Female

A comparison of simplified methods for lipoprotein quantification using the analytic ultracentrifuge as a standard.

Two simplified methods for quantitative lipoprotein analysis have been calibrated and compared with each other using analytic ultracentrifugation as a standard reference procedure. The first method was the Friedewald procedure and the second was an automated agarose gel electrophoresis system. Both procedures offer comparable quantitative lipoprotein analysis with potential for large scale screening purposes at low cost ($4.00-$5.00 per analysis). There were advantages and limitations to both procedures. The Friedwald procedure can be used on frozen sera but requires 3 ml sera. In contrast, the electrophoresis system must be used with fresh serum but requires only 50 mul serum and the electrophoretic slides may be quantitatively analyzed several years retrospectively.

Adult

Repeated scoring with the adult appendicitis score improves the sensitivity and the specificity of appendicitis diagnosis in patients with early equivocal signs of appendicitis: a secondary analysis.

PURPOSE: The utilization of computed tomography in the early stage of acute appendicitis may result in overdiagnosis and unnecessarily expose patients to ionising radiation. The Adult Appendicitis Score (AAS) can be used to select patients for imaging. Observation and re-scoring in the DIAMOND trial reduced the need for imaging. Now, we wanted to determine if the change in AAS (&#x2206;AAS) can serve as a diagnostic tool to select patients for imaging even more precisely. METHODS: Eighty-eight patients with early equivocal appendicitis participated in the observation arm of the DIAMOND trial. The data for these patients were reanalysed, and &#x2206;AAS during the observation was calculated. The baseline AAS, final AAS, and the change in C-reactive protein (&#x2206;CRP) were selected as reference standards. RESULTS: Eighty-three patients with complete data were included in the analysis. The AUROC (Area Under the Receiver Operating Characteristic) values are as follows: &#x2206;AAS, 0.932 (95% CI 0.868-0.996); baseline AAS, 0.629 (95% CI 0.498-0.760); final AAS, 0.936 (95% CI 0.886-0.987); and &#x2206;CRP, 0.796 (95% CI 0.696-0.897). Using receiver operating characteristic curves, we established the thresholds for low (AAS&#x2009;&#x2264;&#x2009;-2), intermediate (AAS -1 to 0), and high (AAS&#x2009;&#x2265;&#x2009;1) probability of appendicitis. The negative predictive value for the low-probability group and the positive predictive value for the high-probability group concerning acute appendicitis were 97% and 94%, respectively. CONCLUSION: Patients with equivocal signs of appendicitis may benefit from short observation and the calculation of &#x2206;AAS to reduce overdiagnosis and exposure to excessive imaging. REGISTRATION: The DIAMOND trial was officially registered on ClinicalTrials.gov (NCT02742402) on April 13, 2016.

Adult