Small Aliquots Research Articles

Simultaneous Determination of Aerosol Inorganic and Organic Nitrogen by Thermal Evolution and Chemiluminescence Detection.

Inorganic nitrogen (IN) and organic nitrogen (ON) molecules constitute a significant part of atmospheric aerosol. Unlike IN, the total ON quantity remains largely unquantified due to a lack of a simple and direct measurement method. This analytical deficiency hinders the quantitative assessment of the various environmental and health effect impacts by aerosol ON. In this work, we developed an analyzer system that utilizes programmed thermal evolution of carbonaceous and nitrogenous aerosols and chemiluminescence detection coupled with the multivariate curve resolution data treatment to achieve simultaneous quantification of IN and ON. The system is capable of detecting IN and ON as low as 96 ng N per sample on a small filter aliquot (1 cm2) without any pretreatment. This method breakthrough opens the door to quantifying an important pool of aerosol N that was analytically inaccessible in the past and holds the promise to quantifying IN and ON in other environmental samples. As a demonstration, quantification of aerosol ON at an urban site in Hong Kong, China, in samples spanning over a year reveals ON constituting a significant fraction (9-52%) of the total aerosol nitrogen and having major source origins in both secondary formation and primary emissions.

Clinical Application of the Fluid Challenge Approach in Goal-Directed Fluid Therapy: What Can We Learn From Human Studies?

Resuscitative fluid therapy aims to increase stroke volume (SV) and cardiac output (CO) and restore/improve tissue oxygen delivery in patients with circulatory failure. In individualized goal-directed fluid therapy (GDFT), fluids are titrated based on the assessment of responsiveness status (i.e., the ability of an individual to increase SV and CO in response to volume expansion). Fluid administration may increase venous return, SV and CO, but these effects may not be predictable in the clinical setting. The fluid challenge (FC) approach, which consists on the intravenous administration of small aliquots of fluids, over a relatively short period of time, to test if a patient has a preload reserve (i.e., the relative position on the Frank-Starling curve), has been used to guide fluid administration in critically ill humans. In responders to volume expansion (defined as individuals where SV or CO increases ≥10–15% from pre FC values), FC administration is repeated until the individual no longer presents a preload reserve (i.e., until increases in SV or CO are <10–15% from values preceding each FC) or until other signs of shock are resolved (e.g., hypotension). Even with the most recent technological developments, reliable and practical measurement of the response variable (SV or CO changes induced by a FC) has posed a challenge in GDFT. Among the methods used to evaluate fluid responsiveness in the human medical field, measurement of aortic flow velocity time integral by point-of-care echocardiography has been implemented as a surrogate of SV changes induced by a FC and seems a promising non-invasive tool to guide FC administration in animals with signs of circulatory failure. This narrative review discusses the development of GDFT based on the FC approach and the response variables used to assess fluid responsiveness status in humans and animals, aiming to open new perspectives on the application of this concept to the veterinary field.

Isolation of Quartz Grains for Optically Stimulated Luminescence (OSL) Dating of Quaternary Sediments for Paleoenvironmental Research.

Optically stimulated luminescence (OSL) dating quantifies the time since mineral grains were deposited and shielded from additional light or heat exposure, which effectively resets the luminescence clock. The systematics of OSL dating is based on the dosimetric properties of common minerals, like quartz and feldspar. The acquired luminescence with exposure to natural ionizing radiation after burial provides a depositional age for many Quaternary sedimentary systems, spanning the past 0.5 Ma. This contribution details the procedures for separating pure quartz grains of a known range of particle sizes to facilitate luminescence analysis with small or single grain aliquots. Specifically, protocols are given for the needed data and interpretations for effective OSL dating of terrestrial sediment cores or sample tubes from exposures. These cores, 5-20 m long in 1.2 m sections, are split lengthwise and crown-cut leaving 80% of core volume undisturbed, which facilitates sampling of light-protected sediment for OSL dating deep within the core. Sediment samples are then subjected to a series of physical separations to obtain a certain grain-size interval (e.g., 150-250 µm). Magnetic minerals are removed in wet and dry states using magnets. A series of chemical digestions starts with soaking in H2O2 to remove organic matter, followed by HCl exposure to remove carbonate minerals, followed by density separation. Subsequently, grains are soaked in HF for 80 min and after in HCl to render solely quartz grains. The mineralogic purity (>99%) of the quartz extract is quantified with grain petrographic assessment and Raman spectroscopy. Repeating this quartz isolation procedure may be necessary with sediment that contains <15% quartz grains. Excitation of the purified quartz grains by LED-derived blue and IR light allows calculations of the fast and IR depletion ratios, which are metrics to assess the dominance of luminescence emissions from quartz.

Sensitive Measurement of Drug-Target Engagement by a Cellular Thermal Shift Assay with Multiplex Proximity Extension Readout.

The ability to monitor target engagement in cellular contexts is a key for successful drug discovery and also valuable in clinical routine. A cellular thermal shift assay (CETSA) provides realistic information about drug binding in cells and tissues, revealing drug-target engagement in clinically relevant samples. The CETSA combined with mass spectrometry (MS) detection can be applied in the early hit identification phase to generate target engagement data for large sets of proteins. However, the analysis is slow, requires substantial amounts of the sample material, and often misses proteins of specific interest. Here, we combined the CETSA and the multiplex proximity extension assay (PEA) for analysis of target engagement of a set of 67 proteins from small amounts of the sample material treated with kinase inhibitors. The results were concordant with the corresponding analyses read out via MS. Our approach allows analyses of large numbers of specific target proteins at high sensitivity in limited sample aliquots. Highly sensitive multiplex CETSA-PEA assays are therefore promising for monitoring drug-target engagement in small sample aliquots in the course of drug development and potentially in clinical settings.

Reduced graphene oxide impregnated in TiO2 for photodegradation of dyes monitored in UV-LED mini-reactor

The photocatalysts of rGO supported on TiO2 were obtained in situ by a sequence of an improved Hummers' method and an eco-friendly hydrothermal procedure. The XRD diffractograms did not present the signals corresponding with the rGO. However, they were detected in micro-Raman because of the characterized signals of D and G bands observed. From TEM, the rGO sheets were visualized covering the surface of TiO2. The photocatalytic experiments were conducted in a real-time monitoring UV-LED mini-reactor (3.0 mL, 5 W and λmax of 365 nm) with small aliquots of sample and catalysts. The catalysts were employed to photodegrade the organic dyes: methylene blue (MB) and tartrazine (TAR), as models or recalcitrant pollutants. The photocatalytic kinetic curves were described by first order kinetic model. Both dyes were completely degraded by all photocatalysts with up to 250 min of irradiation. All supported rGOTiO2 catalysts presented faster kinetics than the TiO2. For MB degradation, 1rGOTiO2 was 6× faster and for TAR degradation 1rGOTiO2 was 3× faster than TiO2. The main action of 1rGOTiO2 was attributed to the better charge transfer by the higher sp2 carbon domains. This result indicates that rGOTiO2 are promising photocatalysts for the degradation of recalcitrant organic pollutants.

Rapid and simultaneous determination of histidine metabolism intermediates in human and mouse microbiota and biomatrices.

Histidine metabolism is a key pathway physiologically involved in satiety, recognition memory, skin, and neural protection and allergic diseases. Microbiologically-produced imidazole propionate induces type II diabetes and interferes with glucose lowering drugs. Despite their determinant health implications, no single method simultaneously assesses histidine metabolites in urine, feces, and microbiota. The aim of this study was to develop a simple, rapid, and sensitive method for the determination of histidine and its major bioactive metabolites histamine, N-acetylhistamine, imidazole-4-acetate, cis-urocanate, trans-urocanate, glutamate and imidazole propionate, using ultrahigh-performance liquid chromatography with electrospray ionization tandem mass spectrometry. An innovative simple extraction method from small aliquots of human and mice urine, feces and microbial cell extracts was coupled to separation in a 6.5min chromatographic run. The successful performance allowed accurate and precise quantification of all metabolites in mouse feces, suggesting broad exchange of histidine metabolites between the gut and mice. Higher urine histamine, histamine to histidine ratio, and imidazole-4-acetate pointed to an underlying inflammatory or allergic process in mice compared to human subjects. N-acetylhistamine and imidazole propionate were detected in human and mouse feces, confirming its origin from gut microbial metabolism. Our novel and robust analytical method captured histidine metabolism in a single assay that will facilitate broad and deep histidine metabolic phenotyping assessing the impact of microbiota on host health in large-scale human observational and interventional studies.

A simple approach for effective shearing and reliable concentration measurement of ultra-high-molecular-weight DNA.

Pipetting and concentration measurement of viscous ultra-high-molecular-weight (UHMW) DNA samples is challenging and often highly imprecise. Effective guidelines for handling UHMW samples are missing in the field. Herein, a simple and low-cost workflow is presented that enables accurate pipetting and reliable concentration measurement. Central to the workflow is the shearing of representative small aliquots of UHMW DNA samples to a fragmentsize <150kb by vortexing them for 1minwith a glass bead in a round-bottomed 2-ml tube. Additionally, a solution is provided for accurate quantitation of high-molecular-weight DNA with fluorometric (Qubit[Thermo Fisher Scientific, MA, USA]) methods by using an appropriate genomic DNA standard, resulting in values that match spectrophotometric (Nanodrop[Thermo Fisher Scientific]) optical densityreadings.

Technical note: On the reliability of laboratory beta-source calibration for luminescence dating

Abstract. The dose rate of the 90Sr / 90Y beta source used in most luminescence readers is a laboratory key parameter. There is a well-established body of knowledge about parameters controlling accuracy and precision of the calibration value but some hard-to-explain inconsistencies still exist. Here, we have investigated the impact of grain size, aliquot size and irradiation geometry on the resulting calibration value through experiments and simulations. The resulting data indicate that the dose rate of an individual beta source results from the interplay of a number of parameters, most of which are well established by previous studies. Our study provides evidence for the impact of aliquot size on the absorbed dose in particular for grain sizes of 50–200 µm. For this grain-size fraction, the absorbed dose is enhanced by ∼ 10 %–20 % as aliquot size decreases due to the radial increase of dose rate towards the centre of the aliquot. The enhancement is most variable for 50–100 µm grains mounted as aliquots of &lt; 8 mm size. The enhancement is reversed when large grains are mounted as small aliquots due to the edge effect by which the dose induced by backscattered electrons is reduced. While the build-up of charge dictates the increase of absorbed dose with the increase of grain size, this principle becomes more variable with changing irradiation geometry. We conclude that future calibration samples should consist of subsamples composed of small, medium, large and very large quartz grains, each obtaining several gamma doses. The calibration value measured with small, medium and large aliquots is then obtained from the inverse slope of the fitted line, not from a single data point. In this way, all possible irradiation geometries of an individual beta source are covered, and the precision of the calibration is improved.

A Simple Sensor System for Onsite Monitoring of O2 in Vacuum-Packed Meats during the Shelf Life.

Vacuum packaging (VP) is used to reduce exposure of retail meat samples to ambient oxygen (O2) and preserve their quality. A simple sensor system produced from commercial components is described, which allows for non-destructive monitoring of the O2 concentration in VP raw meat samples. Disposable O2 sensor inserts were produced by spotting small aliquots of the cocktail of the Pt–benzoporphyrin dye and polystyrene in ethyl acetate onto pieces of a PVDF membrane and allowing them to air-dry. These sensor dots were placed on top of the beef cuts and vacuum-packed. A handheld reader, FirestinGO2, was used to read nondestructively the sensor phase shift signals (dphi°) and relate them to the O2 levels in packs (kPa or %). The system was validated under industrial settings at a meat processing plant to monitor O2 in VP meat over nine weeks of shelf life storage. The dphi° readings from individual batch-calibrated sensors were converted into the O2 concentration by applying the following calibration equation: O2 (%) = 0.034 * dphi°2 − 3.413 * dphi° + 85.02. In the VP meat samples, the O2 levels were seen to range between 0.12% and 0.27%, with the sensor dphi signals ranging from 44.03° to 56.02°. The DIY sensor system demonstrated ease of use on-site, fast measurement time, high sample throughput, low cost and flexibility.

DOES THE PLATELET CONCENTRATION IN PLATELET RICH PLASMA INFLUENCE THE OUTCOMES OF PRIMARY KNEE OSTEOARTHRITIS?

Background: Growth factors in platelets have been extensively studied and were reported to be used to stimulate cartilage regeneration in osteoarthritis (OA). Objective: This study aimed to observe the influence of platelet concentration in platelet rich plasma (PRP) on the outcomes of primary knee OA. Methods: Eighty-nine patients undergoing PRP injection in unilateral primary knee OA were assessed using the Western Ontario and McMaster Universities Arthritis Index (WOMAC) questionnaire and visual analog scale (VAS) before intervention at 3 weeks, 3 months, 6 months and 12 months after treatment. A small aliquot of PRP was sent for bacteriologic examination and evaluation of the platelet count. A student t-test was conducted to compare WOMAC and VAS score among patients before PRP injection (baseline) and at each follow-up. The platelet count and their influence on outcomes were also analyzed using Pearson`s correlation coefficient. Results: Statistically significant differences were observed in the WOMAC score between baseline (M=47.08, SD=8.50) and 3 weeks (M=20.37, SD= 10.09, p&lt; 0.001), 3 months (M= 23.24, SD= 11.39, p&lt;0.001), 6 months (M= 29.89, SD=14.95, p &lt;0.001), and final follow-up at 12 months (M= 27.78, SD= 16.56, p&lt;0.001). Also a significant difference was observed in VAS between baseline (M=69.02, SD= 9.58) and 3 weeks (M= 36.23, SD= 15.72, p &lt;0.001), 3 months (M= 37.04, SD= 17.30, p &lt;0.001), 6 months (M= 42.58, SD=22.15, p &lt;0.001) and 12 months (M=39.15, SD= 23.96, p &lt;0.001). The mean platelet count in PRP injection was 1000.66x103platelets/mL (402x103platelets/ml to 1630x103platelets/mL). Positive correlations were discovered between the concentration of the platelet and the mean improvement WOMAC scores and VAS at 3 weeks (r =0.31, r=0.40), 3 months (r=0.10, r=0.23), 6 months (r=0.08, r=0.30) and 12 months after intervention (r=0.12, r=0.23), respectively. Conclusion: Higher concentrations of platelets in the PRP had a better effect on outcomes of primary knee OA especially at three weeks after injection.

Regulation of transgelin and GST-pi proteins in the tissues of hamsters exposed to sodium arsenite

Regulation of transgelin and GST-pi proteins in the tissues of hamsters exposed to sodium arsenite

Does and duration of exposure time dependent regulation of proteins in the urinary bladder of hamsters exposed to sodium arsenite

Does and duration of exposure time dependent regulation of proteins in the urinary bladder of hamsters exposed to sodium arsenite

Assessment of TMT Labeling Efficiency in Large-Scale Quantitative Proteomics: The Critical Effect of Sample pH.

Isobaric labeling via tandem mass tag (TMT) reagents enables sample multiplexing prior to LC–MS/MS, facilitating high-throughput large-scale quantitative proteomics. Consistent and efficient labeling reactions are essential to achieve robust quantification; therefore, embedded in our clinical proteomic protocol is a quality control (QC) sample that contains a small aliquot from each sample within a TMT set, referred to as “Mixing QC.” This Mixing QC enables the detection of TMT labeling issues by LC–MS/MS before combining the full samples to allow for salvaging of poor TMT labeling reactions. While TMT labeling is a valuable tool, factors leading to poor reactions are not fully studied. We observed that relabeling does not necessarily rescue TMT reactions and that peptide samples sometimes remained acidic after resuspending in 50 mM HEPES buffer (pH 8.5), which coincided with low labeling efficiency (LE) and relatively low median reporter ion intensities (MRIIs). To obtain a more resilient TMT labeling procedure, we investigated LE, reporter ion missingness, the ratio of mean TMT set MRII to individual channel MRII, and the distribution of log 2 reporter ion ratios of Mixing QC samples. We discovered that sample pH is a critical factor in LE, and increasing the buffer concentration in poorly labeled samples before relabeling resulted in the successful rescue of TMT labeling reactions. Moreover, resuspending peptides in 500 mM HEPES buffer for TMT labeling resulted in consistently higher LE and lower missing data. By better controlling the sample pH for labeling and implementing multiple methods for assessing labeling quality before combining samples, we demonstrate that robust TMT labeling for large-scale quantitative studies is achievable.

Mesenchymal stromal cell treatment of donor kidneys during ex vivo normothermic machine perfusion: A porcine renal autotransplantation study.

Normothermic machine perfusion (NMP) of injured kidneys offers the opportunity for interventions to metabolically active organs prior to transplantation. Mesenchymal stromal cells (MSCs) can exert regenerative and anti-inflammatory effects in ischemia-reperfusion injury. The aims of this study were to evaluate the safety and feasibility of MSC treatment of kidneys during NMP using a porcine autotransplantation model, and examine potential MSC treatment-associated kidney improvements up to 14days posttransplant. After 75min of kidney warm ischemia, four experimental groups of n=7 underwent 14h of oxygenated hypothermic machine perfusion. In three groups this was followed by 240min of NMP with infusion of vehicle, 10million porcine, or 10million human adipose-derived MSCs. All kidneys were autotransplanted after contralateral nephrectomy. MSC treatment did not affect perfusion hemodynamics during NMP or cause adverse effects at reperfusion, with 100% animal survival. MSCs did not affect plasma creatinine, glomerular filtration rate, neutrophil gelatinase-associated lipocalin concentrations or kidney damage assessed by histology during the 14days, and MSCs retention was demonstrated in renal cortex. Infusing MSCs during ex vivo NMP of porcine kidneys was safe and feasible. Within the short posttransplant follow-up period, no beneficial effects of ex vivo MSC therapy could be demonstrated.

Allometric equations to estimate aboveground biomass of Dalbergia cearensis species in the Brazilian seasonally dry tropical forest

The semiarid savannas, in particular the Caatinga Biome, are important components of the terrestrial carbon cycle in terms of global carbon storage. Despite this importance, little information is available regarding biomass production potential of caatinga species. The objective of the study was to develop allometric equations to estimate aboveground biomass of Dalbergia cearensis Ducke trees. The semi-destructive method was used to collect samples in which the diameter at breast height (D), wood density (ρ), and tree height (H) were the aboveground dry biomass (AGB) estimation variables. The research was conducted in three different localities of the semiarid region in the state of Ceará, Brazil. Fresh biomass measurement was divided in two parts: trimmed fresh biomass and untrimmed fresh biomass. Measurements were taken for 30 trees. Height, trunk diameter, and branch diameters were measured for each individual. Leaf and small branch aliquots were taken. The volume of branch aliquots was utilized to calculate wood density. Tree dry biomass was obtained by adding trimmed dry biomass and untrimmed dry biomass (AGB, in kg). In all, eight allometric equations were developed for D. cearensis and tested for goodness-of-fit statistics. The best allometric equations were selected based on each model’s performance statistics (R2-adj, RSE, and AIC). Of the eight regression models developed and tested to estimate aboveground biomass of Dalbergia cearensis trees, six attained acceptable performance statistics. The regression model for D. cearensis that utilized the single compound predictive variables DH (model 1) was the most robust, followed by the model using single compound variables ρDH (model 2), and multiple variables D + H (model 3). The methodology applied in this study can be adopted to estimate biomass and volume of a broader set of other species from the Brazilian seasonally dry tropical forest.

Design and Computational Modeling of Spiral Microfluidic Channel for Sorting and Separating the Biomolecules

Background: Microfluidic technologies are a very challenging area today in the field of biomolecule analysis. This has become feasible with the today’s advanced technologies by designing and fabricating the microfluidic channel. Materials and Methods: Initially, microfluidic channels are used to separate large molecules, where the molecular dimension of the fluidic filter is greater than the gap size. In this work, separation of biomolecules (like RBC, WBC and platelets) that are smaller than the microfluidic filter gap size is demonstrated. Results and Conclusion: Due to the curvilinear nature of the spiral, there exists two vortices called dean vortices within the channel and this is influenced by dean flow, centrifugal flow and tubular pinch effect. While flowing a small aliquot of blood in the channel, due to these three effects, molecules attain equilibrium position at one point. The position of equilibrium will be different for different sized biomolecules and this varies with different input velocities. Conclusion: The obtained computational modeling results show how the equilibrium positions influence the separation efficiency of biomolecules in passive based microfluidic filter. Compared with the traditional random nanoporous materials such as gel or polymer monolith, spiral based microfluidic channels can be made precisely to have a pre-determined loop count and Dean Flow number (De).

SISTEMA DE BAIXO CUSTO PARA EXECUÇÃO E MONITORAMENTO ON-LINE DE REAÇÕES FOTOCATALÍTICAS: APLICAÇÃO EM REDUÇÃO DE NITRO-FENOL

LOW-COST SYSTEM FOR ONLINE MONITORING OF PHOTOCALYTICAL REACTIONS: APPLICATION IN NITROPHENOL. There is a global concern with pollutants and organic contaminants and efficient and inexpensive methods are sought for the treatment of these toxic products. Generally, large volume reactors and the use of expensive equipment are required to monitor a photoreaction. The present work proposes the construction of a colorimetric equipment, which allows the online monitoring of photoreactions, using low-cost materials that allow the analysis from small aliquots. To show the applicability of the system, we studied the reduction reaction of 4-nitrophenol (4NP) with NaBH4. The impedimetric sensor enabled to construct the kinetic curve of the decrease in the chromophore band. It was possible to observe that the 4NP reduction reaction depends on: (i) chromophore concentration; (ii) stability of the reducing agent and; (iii) repulsion between the anions. The system developed here opens doors, as it can be used using a photocatalyst in the form of film, and it has also proved to be an inexpensive alternative for both research and teaching laboratories, as it allows the execution of chemical physics practices such as kinetic monitoring of reactions and mathematical adjustment of the curves.

High-Throughput Whole-Plate Imaging of Cells for Multiple Biological Applications.

Advanced multipurpose cell imaging systems along with integrated rapid quantitation software can enhance and expedite cancer cell culture studies in a variety of applications. Though accurate cell culture studies are an important and necessary component of nearly all cancer biomarker detection and therapy studies, the methods we currently use are of low-throughput, time consuming, and lack accuracy. Hence, it is important to improve several features of the assays to increase the accuracy of their quantitative outputs in most studies. In general, we perform cell culture analysis semimanually by counting a small aliquot of suspended cells using a hemocytometer or viewing a small area of cells on a plate using a bright-field microscope, and then extrapolate the counts or observations to estimate the values for the total numbers of cells. The fundamental problem with this process lies in using techniques, such as extrapolation, which inherently introduces intrasample variability while collecting the cells by enzymatic trypsinization for these assays that are affecting cell growth and other downstream assessments. Fluorescence (FL) microscopy-based assays are also used to image and count cells for various applications, including cell viability, proliferation, apoptosis, cell death, transfection efficiency, protein expression, stem cell properties, colony formation, cytotoxicity, drug dose-response, and treatment efficacy studies. These methods are not optimal for many researches, as they require real-time visualization under a microscope plus manual analysis to determine the final results. Owing to long exposure times for cells under fluorescent light of a microscope, the cells may be exposed to suboptimal conditions that affect cell growth, and with occasional photobleaching of the expressed FL probes. Alternatively, the use of cell imaging systems that integrate both advanced bright-field and FL imaging for cell counting and quantification can be useful. In this protocol, we discuss the advantages of a high-throughput cell imaging system using a whole-plate imaging format when used in various bioimaging studies by highlighting a few applications of the system. The system is designed to fundamentally improve the accuracy and time of cell culture analysis while also allowing us to perform the assay without trypsinization, thus avoiding the need to replicate multiple wells for monitoring cell growth over time.

New loading method for high precision Sm isotope analysis of nuclear materials using thermal ionization mass spectrometry

This work demonstrates an analytical protocol for high precision Sm isotope analysis by thermal ionization mass spectrometry (TIMS) using a Pt activator. The method permits precise measurements of small aliquots (1–20 ng) of Sm on single Re filament using a modified static-total evaporation technique. This study represents the first attempt to use such protocols for Sm isotope analyses while reducing the loading size of Sm for TIMS. The method could potentially be deployed to study geological, meteorites and lunar samples containing low Sm concentrations, to monitor neutron irradiation exposure based on 149,150Sm, or to measure Sm isotopic composition in other types of nuclear samples.

Establishing a common standardised growth curve for single-aliquot OSL dating of quartz from sediments in the Jilantai area of North China

Abstract Establishing a common standardised growth curve (SGC) can substantially reduce the instrumental time for equivalent-dose (De) measurements in optically stimulated luminescence (OSL) dating. Several studies have indicated that different samples have different dose–response curves (DRCs) and therefore that it is difficult to construct a common SGC, although an SGC has been proposed in some cases. In this study, our aims were to construct a regional SGC based on small aliquots of sedimentary quartz from more than 100 samples from different sedimentary environments in the Jilantai Basin in North China and to investigate the applicability of different methods of establishing an SGC for the area. The precision of the De values of aliquots which were obtained using the SGC was compared with those obtained using the single-aliquot regenerative (SAR) protocol. Our results indicate the following: (1) for establishing an SGC using the regenerative normalisation (Re-SGC) method, selecting a suitable re-normalisation dose that is close to double the characteristic saturation dose, 2D0, can reduce the inter-aliquot/inter-sample variation in the form of DRCs within a larger dose range. (2) A common regional SGC can be established for the Jilantai area using the Re-SGC and least-squares normalisation (LS-SGC) methods, which provides reliable dating results within the 200 Gy De range.