Evaluating the efficacy of the VAMS Mitra microsampler for whole blood trace element analysis

ABSTRACT

Background

Volumetric Absorptive Microsampling (VAMS) is a blood sample collection method proposed as an alternative to venipuncture for metals/elements biomonitoring. However, the microsampler background concentration of metals and small blood volume remains critical limitations, particularly for metals or trace element analysis in environmental health and epidemiological research.

Materials & methods

Trace element analysis was performed to measure metal concentration in blood samples collected via VAMS and venipuncture using Inductively Coupled Plasma Mass Spectrometry (ICP-MS). VAMS blanks showed elevated background concentration, and cleaning was attempted using a multi-step cleaning protocol. Detection limits and efficacy of background concentration reduction were evaluated.

Results

Initial analysis showed elevated background metal concentrations in the VAMS blank samplers, at or exceeding levels found in venous blood samples. VAMS blank with background concentration post-cleaning result indicated reductions in concentration for some metals; however, the concentration for most detected metals remained persistent.

Conclusions

The efficacy of VAMS in environmental health and epidemiological biomarker research demonstrates both the promising potential and limitations, and the effectiveness of a rigorous cleaning protocol in reducing or eliminating background metal concentrations on microsampler tips.

1. Introduction

Microsampling techniques have transformed human biomonitoring by streamlining the processes of sample collection, storage, and transportation [1]. The introduction of alternative, less invasive blood sampling methods compared with traditional venipuncture dates back to 1913, when Ivar Bang first utilized the dried blood spot (DBS) technique for monitoring glucose levels [2]. By 1963, Guthrie and Susi demonstrated its effectiveness in newborns’ phenylketonuria screening and diagnosis [3,4]. Using a safety lancet, the DBS technique involves placing a drop of blood on a filter card, usually drawn from a finger, heel, or toe. The typical extraction from these spots ranges between 3 and 12 mm in diameter, corresponding to about 80 μL of blood [5,6]. DBS evolved to include a wide array of applications beyond its initial use, encompassing fields like toxicokinetics, epidemiology, and forensic toxicology. Its simplicity, speed, and minimal invasiveness make it preferable over traditional venous blood draw methods while significantly reducing discomfort and the risk of transmitting blood-borne diseases [7,8]. The technique involves small blood volumes and simplifies transport and storage, critical advantages in toxicological applications. However, challenges such as accurate blood volume determination, potential background contamination on the filter card, and the need for rigorous control measures remain significant concerns, necessitating multiple tests to ensure accuracy and minimize false positives [9–11].

To address the limitations of traditional DBS sampling, a novel technique known as volumetric absorptive microsampling (VAMS) was developed in 2014 [12,13]. This method features a hydrophilic polymer tip attached to a plastic handle, which consistently and accurately absorbs a predetermined blood volume via wicking, effectively eliminating the influence of hematocrit levels that affected previous sampling methods [2,14]. Since its introduction, VAMS has been applied in over 150 research studies, primarily in clinical and pharmaceutical research, and has been recently used to collect blood samples for trace element analysis [1,13,15–18]. The design of VAMS devices, which can accurately collect specific blood volumes between 10, 20, and 30 µL, allows samples to be stored and transported easily in diverse formats like cartridges and clamshells. VAMS improves upon DBS by offering a more precise blood volume collection that does not depend on hematocrit levels and provides a more user-friendly collection process compared with traditional venipuncture [2,16].

VAMS has been proposed as an alternative blood sampling method compared to the gold-standard venous sampling method for metal biomonitoring in the general population. Blood-based biomarkers are a practical and reliable approach to characterizing exposures to metals and other environmental toxicants [1,12]. Environmental exposures are increasingly recognized as significant factors associated with adverse health outcomes such as neurodevelopmental disorders in children, necessitating mitigation efforts, when possible, along with close monitoring and early intervention. While the etiology of these health effects remains complex and multifaceted, involving both genetic and environmental factors [19,20], the role of environmental pollution, particularly metals such as cadmium (Cd), chromium (Cr), lead (Pb), manganese (Mn), and mercury (Hg), has garnered attention in recent studies [17,21–24]. This growing body of research highlights the importance of identifying and reducing exposures to improve overall health outcomes. These metals contribute to health impacts and processes, such as permanent damage to the central nervous system, cell apoptosis, oxidative stress, chromosomal damage, and brain damage, contributing to neurodevelopmental disorders, including Autism Spectrum Disorder (ASD) [24–28]. It has been recognized that exposure biomarkers highlight the body burden of metals caused by air pollution, water pollution, and diet; thus, it is identified as a highly effective method for measuring exposures and directly detecting pollutants [25,29,30].

Various research have explored the use of VAMS technology for blood metal analysis. A German study by Koutsimpani-Wagner et al. demonstrated its reliability and precision in quantifying mercury levels. Meanwhile, another study in Canada by Breton et al. focused on a broader range of toxic and essential trace elements and highlighted VAMS as a promising alternative sampling procedure. However, the authors indicated that microsampler contamination poses a critical challenge when using volumetric absorptive microsampling (VAMS) for trace element analysis, necessitating an effective cleaning process. Contaminants such as aluminum (Al), Cr, Mn, molybdenum (Mo), and Pb originate from fabrication processes, environmental factors, and handling. To address this, the authors suggested that a cleaning procedure involving a 2% nitric acid wash followed by a Milli-Q water rinse would significantly reduce contamination for most elements. However, persistent contamination in elements like Al, Cr, Mn, and Mo, among others, limits their reliable quantification and highlights the critical need for an advanced and optimized cleaning procedure for their broader adoption in trace metal assessment [18]. Thus, this study aimed to investigate the efficacy of Volumetric Absorptive Microsampling for collecting blood samples to measure metal exposure in children with ASD. Furthermore, we evaluated the effectiveness of the treatment process to eliminate or reduce background concentration on the microsampler tips for sample collection.

2. Materials and methods

2.1. Study design

Blood samples were collected from 22 participants, 3–10 years with ASD, using Volumetric Absorptive Microsampling (VAMS) and standard venipuncture, with 90 µL of blood obtained via VAMS (three sampler tips, each collecting 30 µL) and approximately 3–4 ml via venipuncture. One blank sampler for quality control was selected from each clamshell containing four VAMS samplers, and blood samples were collected on the remaining three VAMS samplers. The samples were stored, respectively, the vacutainers containing venous blood samples were stored in a −80ºC refrigerator, and microsamplers’ clamshell were stored in their desiccating bags on shelves in our laboratory at stable room temperature. Prior to sample collection, written and signed informed consent was obtained from each participant’s parent or legal guardian. Blood samples were collected at Children’s of Alabama Hospital by trained phlebotomists, with a parent or guardian present. This study was reviewed and approved under the University of Alabama at Birmingham IRB protocol (IRB-300009712). Sample collection started in February 2024 and ended in May 2024. Then, the first batch of samples was sent to the laboratory for analysis using biosafe sample transportation guidelines.

2.2. Sample analysis

Blood metal analysis was conducted at the Wisconsin State Laboratory of Hygiene (WSLH) in a Trace Element Clean Lab (TECL). This facility, covering approximately 3,000 square feet, is designed for ultra-low-level elemental analysis and operates as an ISO 6 clean room or better. The TECL suite minimizes metal contamination using HEPA-filtered air, routine monitoring of airborne particles, and acid precleaning of materials that contact samples. Acid handling is conducted in a separate room to avoid instrument corrosion from acid fumes. These stringent protocols, combined with low contamination levels and sensitive instruments, enable low limits of quantitation (LoQ) for elemental analysis.

The samples underwent digestion in acid-washed polypropylene containers using ultra-pure nitric acid and hydrogen peroxide, heated to 85°C. Elemental analysis was performed using a Thermo Element XR magnetic sector Inductively Coupled Plasma Mass Spectrometer (ICP-MS), following the EPA 200.8 analytical method. Various isotopes and resolution settings were employed to measure target elements and confirm the absence of isobaric interferences. Counts were normalized to internal standards (gallium, indium, and bismuth), and results were blank-corrected using batch-specific method blanks. Propagated uncertainty was calculated using the root-sum-square of three components: (a) SF-ICP-MS analysis (standard deviation of triplicate analyses), (b) blank subtraction (standard deviation of four method blanks), and (c) digestion recovery (long-term standard deviation of standard reference materials and spike recoveries).

2.3. Reducing background concentration

Initial tests showed elevated metal backgrounds in the VAMS blank samples, with levels comparable to the participants’ venous blood concentrations. As a result, the VAMS blood samples were not digested but retained for future use. Further investigation focused on reducing metal contamination in the VAMS blanks. A new set of blank microsamplers from a different lot was sent to the Laboratory for metal background analysis. Breton et al. demonstrated a promising method to lower background levels by cleaning the samplers with diluted nitric acid. Building on this approach, the WSLH team implemented more aggressive pre-treatment steps using a new set of microsamplers that were different from the previous lot.

The cleaning process involved pre-wetting the VAMS sampler tips with methanol to enhance acid penetration, followed by sonication in 10% nitric acid and 5% hydrochloric acid. After thorough rinsing and additional sonication in ASTM Type 1 Reagent Water, the samplers were dried under HEPA-filtered air. Their performance was verified to ensure that the cleaning process did not alter the VAMS samplers’ absorption properties. Samplers were weighed to a precision of 0.00001 g, used to absorb water, and reweighed to measure water volume before and after acid treatment. Although the uptake time increased slightly following acid treatment, the certified volume remained unchanged, confirming that the cleaning procedure preserved the functional integrity of the VAMS samplers. The Limit of Detection (LOD) for each procedure is calculated using the method blanks, where LOD = mean blank concentration + 3*standard deviation of blanks (multiplied by any dilution factors). A result is considered above the LoQ when the concentration exceeds 2.5 times its expanded uncertainty. Results falling below the LoQ for both methods were classified as undetected. Given that all participants in our study were healthy children with ASD and, to our knowledge, had not been exposed to toxic elements, the levels of some elements remained undetected in the majority of our participants.

2.4. Statistical analysis

To test the statistical difference in metal background concentration before and after the cleaning process for the blank samplers in both batches, the difference in the calculated mean was analyzed using a one-way ANOVA (Analysis of Variance) followed by Tukey’s HSD post-hoc test. A one-way ANOVA was conducted to test significant differences in metal concentrations across the treatment process, and a post-hoc test was done to show comparisons between each treatment process. Statistical testing was done using SAS (SAS Institute, Inc., Cary, NC) and R 4.4.3 statistical software (R Core Team 2024).

3. Results

3.1. Venous blood vs volumetric absorptive microsampling

In this study, the standard venipuncture and the Volumetric Absorptive Microsampling Microsampler were used to collect blood samples from each participant. Utilizing the samples, the metals/elements analyzed included antimony (Sb), Cd, Cr, Hg, Mn, nickel (Ni), tin (Sn), and vanadium (V). The concentration measurement indicates a summary of the analyzed metals. Our initial analysis showed that metal concentrations in blood samples collected via VAMS samplers were near or exceeded the venous blood concentration. The venous sample concentration was uniformly below threshold or reference level for most metals. The concentrations of V, Cr, Mn, Ni, Cd, Sn, Sb, Hg, and Pb were within relatively narrow ranges in venous blood samples. The maximum and minimum concentrations of metals measured in venous blood samples and blank samples are demonstrated in (Table 1). Also, the blank samplers in each clamshell for quality control showed broader concentration ranges, especially for Cr, Ni, Cd, Mn, and Sb. Smaller variations were observed for Hg, V, Sn, and Pb in the blanks. Higher metal concentrations were observed in blank samples compared to venous blood samples, highlighting potential background contamination.

Table 1.

Values of maximum and minimum concentrations of metals in blood samples and initial blank samplers.

Element	Unit	Venous blood (max)	Venous blood (min)	Blank Sample (max)	Blank Sample (min)
Cd	µg/L	0.31	0.04	40.80	9.95
Cr	µg/L	9.18	0.03	173	102
Hg	µg/L	5.58	1.16	3.47	0.90
Mn	µg/L	13.20	2.57	17.70	5.37
Ni	µg/L	3.22	0.21	96.60	50.60
Pb	µg/L	23.30	1.51	32.0	1.12
Sb	µg/L	24.1	5.33	18.00	3.50
Sn	µg/L	2.44	0.57	0.54	0.07
V	µg/L	0.13	0.03	0.80	0.60

µg/L: micrograms per liter.

3.2. Cleaning process/sample treatment

In reference to the initial analysis indicating background concentrations of metals on the blank samplers, the research team proceeded to the cleaning process to investigate the effectiveness of cleaning the microsampler to reduce or eliminate the background metal concentration. As illustrated in (Table 2) showing blank samples from the first batch, washed blank VAMS, and their respective LODs are presented.

Table 2.

Mean metal concentration and detection limit before (pre) and after (post) the cleaning process/treatment.

		Metal Concentration		Detection Limit (LOD)
Element	Unit	Blank VAMS	Washed Blank	Blank VAMS	Washed Blank
Cd	µg/L	0.12	0.14	0.25	0.23
Cr	µg/L	131.50	28.80	130	45.50
Hg	µg/L	0.52	0.12	0.61	1.89
Mn	µg/L	10.95	8.89	21.90	23.30
Ni	µg/L	64.70	27.18	63.90	23.50
Pb	µg/L	17.53	7.65	24.30	4.80
Sb	µg/L	2.44	10.91	3.33	2.20
Sn	µg/L	9.59	3.49	53.70	11.20
V	µg/L	0.69	0.81	1.26	3.57

µg/L: micrograms per liter, LOD: limit of detection.

3.3. Treatment evaluation

The cleaning process included the initial blanks and a new batch of microsamplers. The microsampler tips were cleaned in different stages, utilizing a thorough cleaning process described above to achieve a reduced background concentration in the blank samplers (Table 3).

Table 3.

Metal concentration in washed blank samplers at different stages.

	Unit	Untreated Blank	Treated Water	Methanol/Treated Water	Acid Washed Blank	Reagent (no VAMS)
Element		Mean	Mean	Mean	Mean	Mean
Al	µg/L	888.67	844.0	1139.4	552.7	90.52
Cd	µg/L	0.13	0.13	0.27	0.09	0.14
Cr	µg/L	71.75	25.62	33.74	18.72	1.73
Fe	µg/L	477.17	555.60	811.60	17.52	7.96
Hg	µg/L	0.08	0.16	0.04	0.17	0.08
Mn	µg/L	12.85	9.66	18.55	6.20	0.02
Mo	µg/L	11.24	4.48	6.55	4.48	0.000025
Ni	µg/L	51.07	30.34	63.38	11.74	0.01
Pb	µg/L	9.98	15.93	22.28	1.13	0.01
Sb	µg/L	60.46	1.12	1.25	0.47	0.03
Sn	µg/L	15.35	1.54	5.54	0.04	0.03
V	µg/L	0.61	0.70	0.78	1.16	0.01
Y	µg/L	0.40	0.15	0.88	0.20	0.000002
Zn	µg/L	305.0	87.28	144.86	22.64	15.02

µg/L:micrograms per liter.

The illustration in Figure 1 visually demonstrates the improvement in the blank samplers’ background concentrations of neurotoxic metals following each washing process of the microsampler tips. The background concentration of several metals showed a decreasing trend through the treatment processes. Cr, Mg, and Ni concentrations were high in the untreated blank, steadily decreasing through treated water and acid-washed blanks, reaching their lowest levels in the reagent blank. Similarly, cadmium, tin, and antimony followed this pattern, with significant reductions after treatment and minimal presence in the reagent blank. In contrast, some metals temporarily increased before eventually decreasing. Lead and mercury concentrations increased during the initial treatment process before decreasing when washed with acid-washed. Vanadium also showed an increase in the acid-washed blank but decreased in the reagent blank.

Figure 1.

Demonstrates the improvement in the blank background concentrations of neurotoxic metals following each washing process of the microsampler tips.

In addition to the neurotoxic metals displayed in Table 3, other elements were analyzed during the cleaning process. The broader panel of elements included arsenic (As), aluminum (Al), calcium (Ca), cerium (Ce), dysprosium (Dy), europium (Eu), iron (Fe), lanthanum (La), lutetium (Lu), magnesium (Mg), molybdenum (Mo), neodymium (Nd), niobium (Nb), praseodymium (Pr), rubidium (Rb), selenium (Se), scandium (Sc), samarium (Sm), silver (Ag), strontium (Sr), tantalum (Ta), thallium (Tl), yttrium (Y), and zinc (Zn). On the microsamplers’ tips, Al, Fe, Mo, Y, and Zn showed improvement in decreased background levels using the cleaning process. However, the other elements investigated in the broader panel (e.g., Ca, Ce, Dy, Eu, La, Lu, Mg, Nb, Nd, Pr, Rb, Sc, Sm, Sr, Ta, Tl) appeared intermittently contaminated in the cleaning process.

Some other elements showed improvement in concentrations across the treatment process. Most elements decreased overall, with some temporary increases in the initial process. Al concentration decreased and increased throughout the process but was lowest in the reagent blank. Nickel (Ni) and zinc (Zn) followed a similar pattern, decreasing in treated water, increasing in methanol-treated water, and decreasing substantially in acid-washed and reagent blanks. Iron (Fe) initially increased in treated water and peaked in methanol-treated water before decreasing significantly in the following process. Likewise, yttrium (Y) and molybdenum (Mo) decreased after treatment and were reduced significantly in the reagent blank.

3.4. Statistical analysis

A one-way analysis of variance (ANOVA) was conducted to examine the effect of the treatment process on blank background concentration levels (Table 3). The ANOVA revealed a statistically significant effect of the treatment process on the background concentration (F (4, 40) = 2.93, p-value = 0.0325). Post hoc comparisons using Tukey’s HSD test (α = 0.05, minimum significant difference = 22.895) indicated no statistically significant differences between any pair of treatment process means, as none of the pairwise differences exceeded the minimum significant difference threshold. However, visual inspection of the means suggested trends among individual treatments. The mean concentration for the untreated blanks was highest at 24.6978, methanol/treated water (16.2033), treated water (9.4867), acid wash (4.4133), and reagent solution (0.2289).

4. Discussion

The Volumetric Absorptive Microsampling (VAMS) technique represents a substantial improvement in blood sample collection, by offering precise quantitative bioanalysis, the capability to sample specific volumes of whole blood consistently, and a less invasive method of taking blood from children. VAMS devices are generally precise in their uptake but are not without limitations. One significant limitation is the susceptibility to user error, mainly when samples are collected at home without trained professional supervision. In our study, with trained professionals taking the samples, we observed two blank samples contaminated with blood spots, highlighting the potential issues with mishandling. Moreover, once the blood sample dried, it became difficult to ascertain whether the microsampler tip was underloaded or overloaded. If this occurs, it complicates quality control, can amplify errors when dealing with small volumes, and relative errors due to sampling inconsistencies can be indicated. Another limitation of the VAMS sampler tips is their limited sample volume of 10–90 µL, substantially lower than the typical 100–500 µL volume digested from venous blood samples. This smaller volume increases the relative error from sampling issues and exacerbates background concentration problems. Also, previous studies on medical blood sampling have reported a biologically plausible risk that capillary blood may be relatively diluted by additional plasma or interstitial fluid compared to venous blood; however, a study by Rodríguez-Saldaña, V., & Basu, N. (2022) focused on comparing the use of capillary and venous whole blood to measure toxic metal(loid)s (including the elements measured in our study) found strong correlations for all measured elements between capillary and venous blood samples [31].

The potential of VAMS as a metal/element analysis method could be significantly enhanced by mitigating background contamination and increasing the sample volume. A minimum blood sample volume of 100 µL is preferred for effective analysis, typical for venous blood sampling, which ranges from 100 to 500 µL. However, the limited sample volume obtainable from VAMS sampler tips presents a challenge, particularly as background contamination levels tend to rise when pooling multiple sampler tips to achieve larger volumes. Also, the statistical analysis conducted to test the variability of background concentration in the different treatment processes indicated that the untreated blank samplers had the highest background concentration, and the other treatment processes may have reduced the concentration to some extent, particularly the acid wash. However, the differences between the treatment processes are not largely significant when adjusted for multiple comparisons. Attempting further improvements to the cleaning procedure would require further complications or additional equipment, which defeats the purpose of the simplicity of the VAMS method. Additionally, variations in washing capabilities across different laboratories, especially between academic research labs and health agencies with dedicated surveillance programs, introduce inconsistencies in background contamination. We also saw some evidence of variability between the manufacturers’ lots, further contributing to these inconsistencies, which must be addressed to ensure accurate blood metal measurements.

The cleaning procedure is not guaranteed to be equally effective across labs, and uniformity in background contamination is critical for reliable results. For example, lead is highly ubiquitous; therefore, the overall cleanliness of the laboratory environment itself can contribute to background contamination in the sampler. Some of the earliest clean labs were specifically designed to measure lead accurately, highlighting the importance of stringent contamination controls [32,33]. As Patterson emphasized, accurately determining lead in environmental samples necessitates ‘ultraclean’ laboratory environments to minimize procedural blanks and environmental contamination [33]. A further limitation of the VAMS is the costs associated with cleaning and tracking the sampler tips. The funds needed to cover labor, materials, and overhead can be approximately $70 per batch, though this estimate applies to a batch size of approximately 10–50 sampler tips. Additionally, the extent of contamination at the point of use remains unquantified but could be substantial, given the low sample volumes involved.

VAMS samplers are advantageous for collecting blood samples for research and laboratory settings. VAMS is considered minimally invasive and user-friendly, with benefits including ease of storage, transport, and a cost-effective blood sampling method for research purposes [1,14,18]. For laboratory analysis, microsamplers’ tips are easier to prepare for analysis and handle compared with traditional methods of pipetting and weighing liquid blood due to their design. Likewise, each tip is well labeled with serial numbers, enhancing traceability, which is crucial for systematic analysis.

Our study investigated the advantages of the VAMS method as a modern or advanced blood sample collection method, especially among our sample population, which includes children. The advancing scientific evidence indicates that early exposure to toxic metals is a significant environmental risk factor for neurodevelopmental disorders, particularly autism spectrum disorder (ASD), highlighting the importance of early metal body burden investigation. The study aimed to validate VAMS as a method of collecting blood samples to conduct environmental health and epidemiological biomarker research for multiple metals among children, particularly vulnerable children diagnosed with ASD. We considered VAMS as an accessible blood sampling method to enhance access to a greater proportion of ASD cases, identify risk factors with environmental sources, enable greater participation in future research, and further develop targeted public health interventions.

5. Conclusions

This study highlights the potential use of VAMS as an alternative to traditional venous blood draw and its challenges for collecting blood samples in environmental health research. VAMS offers advantages, particularly as minimally invasive, easy to handle, and its traceability, which make it a suitable alternative for research involving vulnerable populations, such as children with ASD. However, limitations related to blank background metal level, sample volume, susceptibility to contamination, and variability in laboratory procedures must be carefully addressed to ensure reliable and accurate metal analysis.

To fully realize the potential of VAMS, future research should prioritize the ability to increase sample volumes, mitigate contamination, standardize laboratory practices across various settings, and develop comprehensive training protocols for home-based sampling. Despite these challenges, VAMS remains a valuable tool in environmental health and biomarker research, potentially increasing participation among vulnerable populations and supporting more inclusive and accessible public health studies.

Funding Statement

This work was supported by an FCIDD McNulty Award from the Civitan International Research Center at UAB.

Article highlights

• VAMS was evaluated as an alternative to venipuncture in blood metal biomonitoring.

• Background metal concentration on VAMS blood samples and blanks exceeds the venous blood concentration.

• Rigorous cleaning protocol reduced metal background in VAMS blanks.

• VAMS is a valuable potential tool in environmental health and biomarker research.

Author contributions

Oludesola Ogunesan: Conceptualization, methodology, investigation, collection of samples, recruitment, data analysis, visualization, project coordination, writing the original draft, and editing the manuscript. Christa Dahman Zaborske: Blood sample analysis, methodology, resources, review and editing. Martin Shafer: Blood sample analysis, methodology, and reviewing. Cassandra Newsom: Sample collection, review and editing. Kristina Zierold: Reviewing and editing. Jeffrey Wickliffe: Conceptualization, methodology, investigation, reviewing and editing, funding acquisition, and supervision.

All authors revised the manuscript, provided key guidance, and approved the final manuscript.

Disclosure statement

The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.

No writing assistance was utilized in the production of this manuscript.

Reviewer Disclosures

Peer reviewers on this manuscript have no relevant financial or other relationships to disclose.

Ethical declaration

Blood samples were collected under the auspices of IRB approval (IRB-300009712) from children diagnosed with ASD who received research-reliable diagnoses with ADOS-2 at the University of Alabama at Birmingham.

Data availability statement

Data are available from the corresponding author upon request.