The analysis of serum free light chains (FLCs) is clinically relevant for the diagnosis and therapeutic management of clonal plasma cell disorders. This study compares the performance of monoclonal ...and polyclonal FLC κ and λ assays in clinical samples determined in a single academic center.
Serum FLCs were analyzed from 102 patients using the Freelite (Binding Site) and N Latex (Siemens) assays on the BN ProSpec System (Siemens). When available, data for protein electrophoresis, immunofixation, C-reactive protein, and estimated glomerular filtration rate (eGFR) were combined with FLC results to evaluate performance.
Method evaluation showed acceptable imprecision and inaccuracy measures of <4.4% and 12.9%, respectively. Poor agreement between the methods was observed, including constant and proportional bias and poor correlation (Kendall τ, 0.671-0.901). The N Latex assay was not affected by the renal impairment estimated by eGFR, unlike the FLC κ/λ ratio results by the Freelite assay. With the Freelite assay, 98% of putative controls without monoclonal gammopathy (n = 42) showed a κ/λ ratio that was above the median of the standard diagnostic range or renal diagnostic range. A shift toward higher κ/λ ratios was also observed when retrospective data between 2011 and 2017 were compared.
Unlike the Freelite assay, κ/λ ratios analyzed with the N Latex assay were not affected by renal failure. Both methods showed acceptable performances using nephelometry, but they were poorly correlated. A shift toward κ/λ ratios might impair the specificity of borderline increased κ/λ results. This should be considered when interpreting FLC κ and λ results.
•Microarray based MALDI-MS for metabolite and antibody monitoring shows excellent results.•Monoclonal antibody is monitored directly from cell supernatant without prior purification.•The MALDI-MS ...method is significantly faster and cheaper compared to HPLC–UV approaches.
Cell culture process monitoring in monoclonal antibody (mAb) production is essential for efficient process development and process optimization. Currently employed online, at line and offline methods for monitoring productivity as well as process reproducibility have their individual strengths and limitations. Here, we describe a matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS)-based on a microarray for mass spectrometry (MAMS) technology to rapidly monitor a broad panel of analytes, including metabolites and proteins directly from the unpurified cell supernatant or from host cell culture lysates. The antibody titer is determined from the intact antibody mass spectra signal intensity relative to an internal protein standard spiked into the supernatant. The method allows a semi-quantitative determination of light and heavy chains. Intracellular mass profiles for metabolites and proteins can be used to track cellular growth and cell productivity.
Despite the significance of glycoproteins for extracellular matrix assembly in cartilage tissue, little is known about the regulation of the chondrocyte glycophenotype under inflammatory conditions. ...The present study aimed to assess the effect of IL-1β and TNF-α on specific features of the glycophenotype of primary human chondrocytes in vitro. Using LC–MS, we found that both cytokines increased overall sialylation of N- and O-glycans and induced a shift towards α-(2→3)-linked sialic acid residues in chondrocyte glycoproteins. These results were supported by quantitative PCR showing increased expression of α-(2→3) sialyltransferases in treated cells. Moreover, we found that both IL-1β and TNF-α induced a considerable shift from oligomannosidic glycans towards complex-type N-glycans. In contrast, core α-(1→6)-fucosylation of chondrocyte N-glycans was found to be reduced particularly by TNF-α. In summary, inflammatory conditions induce specific alterations of the chondrocyte glycophenotype which might affect cell–matrix interactions or the function of endogenous lectins.
Our study compared sensitivity, specificity, and accuracy of whole-body diffusion-weighted imaging (WB-DWI) using a b-value of 2000 s/mm
with that of the commonly used b-value of 800 s/mm
for ...depiction of active tumor sites in patients with plasma cell diseases. We introduced an ultrahigh b-value to reduce interfering signals from benign and post-therapeutic inactive lesions by suppressing T2-shine-through effects.
The prospective single-center study included patients when they went through a whole-body MRI (WB-MRI) staging or response evaluation procedure. The apparent diffusion coefficient (ADC) and morphologic appearance served as reference for classifying focal lesions on WB-DWI as vital or post-therapeutic. Additionally, we compared our classification with patients' serological markers of disease activity.
One hundred participants (65 ± 10 years, 58 men) underwent WB-DWI between June and October 2019. The detection rate of vital focal lesions was similar for both b-values with a sensitivity of 0.99 using b = 800 s/mm
and 0.98 using b = 2000 s/mm
. By contrast, specificity and accuracy were 0.09 and 0.71 when using a b-value of 800 s/mm
, and 0.96 and 0.98 when using a b-value of 2000 s/mm
, respectively. The difference in specificity and accuracy was statistically significant (p < 0.001).
Using a b-value of 2000 s/mm
significantly improved the specificity of lesion detection with WB-DWI as compared to the commonly used b-value of 800 s/mm
. The high b-value significantly reduced signal intensities of post-therapeutic or benign lesions and provided a significantly more accurate representation of active tumor load.
An electrical measurement is non-linear when it is affected by the applied stimulus, i.e. when the measured phenomenon changes with amplitude. If pinched hysteresis loops can be observed in the ...voltage current representation, the underlying tissue can be classified as a memristor. Several biological memristors have been published, like human skin and apples. However, changes in the polarization impedance of electrodes may also cause pinched hysteresis loops. The question whether the reported biological memristors are real or whether the results just reflect changes in the polarization impedance arises. If the impedance of the measured object is close to or smaller than the polarization impedance of the used electrodes, the latter may dominate the measurement.
In this study, we investigated the non-linear electrical properties of silver/silver chloride electrodes in a sodium chloride solution that has a similar concentration as human sweat and compared these to results from human skin. First of all, we found that silver/silver chloride electrodes in sodium chloride solution can be classified as memristors. However, the currents obtained from the sodium chloride solution are much higher than the currents recorded from human skin and there is a qualitative difference in the pinched hysteresis loops in both cases. We can conclude that the non-linear electrical measurements with silver/silver chloride on human skin are actually dominated by the skin and we can confirm that the human skin memristor really exists.
Early-onset pneumonia (EOP) after out-of-hospital cardiac arrest is frequently observed. Causative factors are loss of airway protection during cardiac arrest, pulmonary contusion, and emergency ...airway management. We assessed the incidence, risk factors, and clinical course of EOP, and evaluated the impact of an early exchange of the prehospitally inserted endotracheal tube (ETT).
In our retrospective analysis we included 104 consecutive subjects admitted to our ICU after out-of-hospital cardiac arrest between 2007 and 2012. All subjects underwent therapeutic hypothermia. We analyzed clinical course, inflammation indicators, Clinical Pulmonary Infection Score, occurrence of EOP, duration of ventilatory support, microbiological findings, and short-term outcome.
Of the 104 subjects, 46.2% received an exchange of ETT directly after hospital admission. Neither ETT exchange nor observed aspiration were associated with elevated CPIS or EOP, nor with proof of microorganisms in respiratory secretions. We found no differences in duration of ventilatory support, P(aO2)/F(IO2), ICU days, or outcome. C-reactive protein was significantly higher in subjects with aspiration (P = .046). Sex, age, smoking status, aspiration, cause of cardiac arrest, first detected heart rhythm, and use of supraglottic airways devices were not associated with EOP. Subjects with EOP had a longer need for ventilatory support (P = .005), higher tracheotomy rate (P = .03), longer ICU stay (P = .005), higher C-reactive protein (P < .001), higher body temperature (P = .003), higher Clinical Pulmonary Infection Score (P < .001), and lower P(aO2)/F(IO2) (P = .008).
The rate of EOP was not significantly influenced by the exchange of the preclinically inserted ETT, but was associated with longer need for mechanical ventilation and ICU stay.
Severe cardiac disorders predispose to central sleep apnoea (CSA). We sought to examine the relationship between severe aortic stenosis, sleep disordered breathing (SDB), and CSA before and after ...transcatheter aortic valve implantation (TAVI).
Twenty-nine patients (81±6 yrs, 41% male, LVEF 48±14%) with severe aortic stenosis and high surgical risk underwent polygraphy before and three months after TAVI. Patients with an apnoea-hypopnoea index (AHI) >5/hr were considered to have SDB. SDB with ≥50% absence of both airflow and ventilatory effort was defined as CSA. Twenty-one of 29 patients (72%) had SDB (6/6/9 mild, moderate, and severe, respectively), 12 (41%) with CSA (0/4/8) and 9 (31%) with obstructive sleep apnoea (7/2/0). There was a strong correlation of CSA with LVEDP before TAVI (r=0.74, p=0.024), but not with LVEF, systolic pulmonary artery pressure or NT pro-BNP. After TAVI, AHI improved significantly, particularly in the CSA group (from 43.5±17.5 to 19.4±12.9/hr, p<0.001). Prevalence and severity of SDB were reduced from 72% to 59% (6/6/9 to 7/8/2 patients), triggered by the significant improvement of CSA.
Patients with severe aortic stenosis display a high prevalence of SDB, particularly of CSA. Definitive treatment with TAVI greatly resolved SDB in the CSA subgroup.
Increasing numbers of bacterial strains being resistant to conventional antibiotics emphasize the urgent need for new antimicrobial agents. One strategy is based on host defence peptides that can be ...found in every organism including humans. We have studied the antimicrobial peptide LF11, derived from the pepsin cleavage product of human lactoferrin, known for its antimicrobial and lipid A-binding activity, and peptide C12LF11, the N-lauryl-derivative of LF11, which has owing to the attached hydrocarbon chain an additional hydrophobic segment. The influence of this hydrocarbon chain on membrane selectivity was studied using model membranes composed of dipalmitoylphosphatidylglycerol (DPPG), mimicking bacterial plasma membranes, and of dipalmitoylphosphatidylcholine (DPPC), a model system for mammalian membranes. A variety of biophysical techniques was applied. Thereby, we found that LF11 did not affect DPPC bilayers and showed only moderate effects on DPPG membranes in accordance with its non-hemolytic and weak antimicrobial activity. In contrast, the introduction of the N-lauryl group caused significant changes in the phase behaviour and lipid chain packing in both model membrane systems. These findings correlate with the in vitro tests on methicillin resistant
S. aureus,
E. coli,
P. aeruginosa and human red blood cells, showing increased biological activity of C12LF11 towards these test organisms. This provides evidence that both electrostatic and hydrophobic interactions are crucial for biological activity of antimicrobial peptides, whereas a certain balance between the two components has to be kept, in order not to loose the specificity for bacterial membranes.
Huntington's disease (HD) is caused by an expanded CAG repeat leading to the synthesis of an aberrant protein and to the formation of polyglutamine (polyQ)-containing inclusions and aggregates. ...Limited information is available concerning the association of neuropathological markers with the development of behavioral markers in HD. Using a previously generated transgenic rat model of HD (tgHD rat), we performed association studies on the time-course of behavioral symptoms (motor function, learning, anxiety) and the appearance of striatal atrophy, 1C2 immunopositive aggregates and polyQ recruitment sites, a precursor to these aggregates. At the age of 1 month, tgHD rats exhibited reduced anxiety and improved motor performance, while at 6 months motor impairments and at 9 months cognitive decline occurred. In contrast, polyQ recruitment sites appeared at around 6–9 months of age, indicating that HD-like behavioral markers preceded the appearance of currently detectable neuropathological markers. Interestingly, numerous punctate sites containing polyQ aggregates were also seen in areas receiving afferents from the densely recruiting regions suggesting either transport of recruitment-competent aggregates to terminal projections where initially 1C2 positive aggregates were formed or different internal properties of neurons in different regions. Furthermore, striatal atrophy was observed at the age of 12 months. Taken together, our findings support the hypothesis of a dynamic process leading to region- and age-specific polyQ recruitment and aggregation. The dissociation of onset between behavioral and neuropathological markers is suggestive of as yet undetected processes, which contribute to the early phenotype of these HD transgenic rats.
PDF
