Pathways to substance use: Examining conduct problems and parenting behaviors from preschool to adolescence.

While many studies have identified risk and protective factors of substance use (SU), few have assessed the reciprocal associations of child conduct problems (CP) and parenting practices and behaviors in the prediction of SU across development. A greater understanding of how these factors relate over time is needed to improve the timing of targeted prevention efforts. This study examined how child CP, parenting behaviors, and parents' own antisocial behavior relate from preschool to adolescence and eventuate in SU. Participants included 706 youth (70.6% male; 89.7% white) enrolled in the Michigan Longitudinal Study. Data from waves 1 (ages 3-5), 2 (ages 6-8), 3 (ages 9-11), 4 (ages 12-14), and 5 (ages 15-17) were included. A random intercept cross-lagged panel model (RI-CLPM) examined reciprocal associations between parenting practices, parents' antisocial behavior, and child CP over time (waves 1-4) and how these factors contribute to adolescent alcohol, cigarette, and marijuana use (wave 5). At the within-person level, negative parenting and parents' own antisocial behavior had a strong influence in late childhood/early adolescence. Only child CP emerged as a significant predictor of SU. Results highlight the importance of early intervention and the potential influence of parenting and child factors throughout development in the prevention of SU.

Individual differences in the development of youth externalizing problems predict a broad range of adult psychosocial outcomes.

This study examined how youth aggressive and delinquent externalizing problem behaviors across childhood and adolescence are connected to consequential psychosocial life outcomes in adulthood. Using data from a longitudinal, high-risk sample (N = 1069) that assessed children and their parents regularly from early childhood (ages 3-5) through adulthood, multilevel growth factors of externalizing behaviors were used to predict adult outcomes (age 24-31), providing a sense of how externalizing problems across development were related to these outcomes via maternal, paternal, teacher, and child report. Findings indicated strong support for the lasting connections between youth externalizing problems with later educational attainment and legal difficulties, spanning informants and enduring beyond other meaningful contributors (i.e., child sex, cognitive ability, parental income and education, parental mental health and relationship quality). Some support was also found, although less consistently, linking externalizing problems and later alcohol use as well as romantic relationship quality. Delinquent/rule-breaking behaviors were often stronger predictors of later outcomes than aggressive behaviors. Taken together, these results indicate the importance of the role youth externalizing behaviors have in adult psychosocial functioning one to two decades later.

Adverse childhood experiences, sleep problems, low self-control, and adolescent delinquency: A longitudinal serial mediation analysis.

Several studies link adverse childhood experiences (ACEs) to delinquency. Yet, developmental sequalae accounting for this association remain unclear, with previous research limited by cross-sectional research designs and investigations of singular mediating processes. To redress these shortcomings, this study examines the longitudinal association between ACEs and delinquency as mediated by both sleep problems and low self-control, two factors which past research implicates as potentially important for understanding how ACEs contribute to antisocial behavior. Data collected from 480 adolescents (71.3% boys; 86.3% White) and their parents participating in the Michigan Longitudinal Study was used to conduct a serial mediation analysis. The association between ACEs (prior to age 11) and delinquency in late adolescence was found to operate indirectly via sleep problems in early adolescence and low self-control in middle adolescence. Nonetheless, a direct association between ACEs and later delinquency remained. Pathways through which ACEs contribute to later delinquency are complex and multiply determined. Findings indicate that early behavioral interventions, including improving sleep and self-control, could reduce later delinquency. Still, more research is needed to identify additional avenues through which the ACEs-delinquency association unfolds across development.

Exposure to Parental Alcohol Use Is Associated with Adolescent Drinking Even When Accounting for Alcohol Exposure of Best Friend and Peers.

AIMS: To further disentangle the role of exposure to drinking of role models (parents, peers, best friends) in the development of young adolescent alcohol use, the current study examined (a) whether parent's alcohol use exposure was associated with alcohol use outcomes among adolescents and (b) whether this association remained significant when including best friend and peer drinking exposure. METHODS: A longitudinal study followed 765 adolescents from the Netherlands over 3 years. Adolescents (45.6% male, Mage = 11.78, standard deviation = 0.49 at baseline) completed questionnaires every 6 months, resulting in seven measurement waves. Adolescents reported their own alcohol use and exposure to parental, best friend and peers drinking. RESULTS: Multilevel regression analyses indicated that parental alcohol use exposure was positively associated with a higher likelihood of adolescent alcohol use in the past 6 months, drinking in the last month and binge drinking in the last month. These associations remained significant when including exposure to peer and best friend's alcohol use, also when controlling for alcohol use at the previous timepoint (i.e. change in drinking). These associations were also consistent for boys and girls. CONCLUSIONS: Throughout early adolescence, parental alcohol exposure matters for their offspring's alcohol use, independently of whether peers or their best friend expose them to alcohol or not. Parental alcohol exposure should be considered in prevention efforts to further decrease the number of adolescents that engage in early alcohol use and binge drinking.

Subtypes of inhibitory and reward activation associated with substance use variation in adolescence: A latent profile analysis of brain imaging data.

The present study identified subgroups based on inhibitory and reward activation, two key neural functions involved in risk-taking behavior, and then tested the extent to which subgroup differences varied by age, sex, behavioral and familial risk, and substance use. Participants were 145 young adults (18-21 years old; 40.0% female) from the Michigan Longitudinal Study. Latent profile analysis (LPA) was used to establish subgroups using task-based brain activations. Demographic and substance use differences between subgroups were then examined in logistic regression analyses. Whole-brain task activations during a functional magnetic resonance imaging go/no-go task and monetary incentive delay task were used to identify beta weights as input for LPA modeling. A four-class model showed the best fit with the data. Subgroups were categorized as: (1) low inhibitory activation/moderate reward activation (39.7%), (2) moderate inhibitory activation/low reward activation (22.7%), (3) moderate inhibitory activation/high reward activation (25.2%), and (4) high inhibitory activation/high reward activation (12.4%). Compared with the other subgroups, Class 2 was older, less likely to have parental alcohol use disorder, and had less alcohol use. Class 4 was the youngest and had greater marijuana use. Classes 1 and 3 did not differ significantly from the other subgroups. These findings demonstrate that LPA applied to brain activations can be used to identify distinct neural profiles that may explain heterogeneity in substance use outcomes and may inform more targeted substance use prevention and intervention efforts.

Lack of Detectable Direct Effects of Silver and Silver Nanoparticles on Mitochondria in Mouse Hepatocytes.

Silver nanoparticles (AgNPs) are well-proven antimicrobial nanomaterials, yet little is elucidated regarding the mechanism underlying cytotoxicity induced by these nanoparticles. Here, we tested the hypothesis that mitochondria are primary intracellular targets of two AgNPs and silver ions in mouse hepatocytes (AML12) cultured in glucose- and galactose-based media. AML12 cells were more sensitive to mitochondrial uncoupling when grown with galactose rather than glucose. However, 24 h treatments with 15 nm AgNPs and 6 nm GA-AgNPs (5 and 10 µg/mL) and AgNO3 (1 and 3 µg/mL), concentrations that resulted in either 10 or 30% cytotoxicity, failed to cause more toxicity to AML12 cells grown on galactose than glucose. Furthermore, colocalization analysis and subcellular Ag quantification did not show any enrichment of silver content in mitochondria in either medium. Finally, the effects of the same exposures on mitochondrial respiration were mild or undetectable, a result inconsistent with mitochondrial toxicity causing cell death. Our results suggest that neither ionic Ag nor the AgNPs that we tested specifically target mitochondria and are inconsistent with mitochondrial dysfunction being the primary cause of cell death after Ag exposure under these conditions.

Neural correlates of inhibitory control in youth with symptoms of food addiction.

Prior research has found that food addiction is associated with reward-related neural differences, but research has yet to examine whether there are also neural differences in inhibitory control. This may be particularly relevant during adolescence as it is a key developmental period where difficulties in inhibitory control are more prevalent. The Yale Food Addiction Scale is a self-report questionnaire that applies substance use disorder diagnostic criteria to certain foods that has also been adapted for children. Here we investigate the association between addictive-like eating and brain functioning during inhibitory control in youth. Seventy-six right-handed participants 8.2-17.8 years (44 male) were recruited. Participants performed a go/no-go task during functional magnetic resonance imaging and completed the Yale Food Addiction Scale for Children, after which they were categorized into two groups according to their scores (No Symptom Group = 0; YFAS-C Group: score ≥ 1). Inhibitory control was probed with a contrast of correct no-go versus go trials. An independent-samples t-test comparing groups revealed a significant difference in three primary clusters, all exclusively in the left hemisphere (No Symptom Group > YFAS-C Group): middle temporal gyrus/occipital gyrus, precuneus/calcarine sulcus, and inferior frontal gyrus. Specifically, the YFAS-C Group showed deactivation in all three clusters. Adolescents who endorse food addiction appear to show hypo-activation in response to the inhibitory control portion of a go/no-go task, which suggests possible inhibitory control difficulties.

Natural Disasters in the Americas, Dialysis Patients, and Implications for Emergency Planning: A Systematic Review.

INTRODUCTION: Natural hazards are elements of the physical environment caused by forces extraneous to human intervention and may be harmful to human beings. Natural hazards, such as weather events, can lead to natural disasters, which are serious societal disruptions that can disrupt dialysis provision, a life-threatening event for dialysis-dependent people. The adverse outcomes associated with missed dialysis sessions are likely exacerbated in island settings, where health care resources and emergency procedures are limited. The effect of natural disasters on dialysis patients living in geographically vulnerable areas such as the Cayman Islands is largely understudied. To inform predisaster interventions, we systematically reviewed studies examining the effects of disasters on dialysis patients and discussed the implications for emergency preparedness in the Cayman Islands. METHODS: Two reviewers independently screened 434 titles and abstracts from PubMed, Scopus, CINAHL, and Cochrane Library. We included studies if they were original research articles published in English from 2009 to 2019 and conducted in the Americas. RESULTS: Our search yielded 15 relevant articles, which we included in the final analysis. Results showed that disasters have both direct and indirect effects on dialysis patients. Lack of electricity, clean water, and transportation, and closure of dialysis centers can disrupt dialysis care, lead to missed dialysis sessions, and increase the number of hospitalizations and use of the emergency department. Additionally, disasters can exacerbate depression and lead to posttraumatic stress disorder among dialysis patients. CONCLUSION: To our knowledge, this systematic review is the first study that presents a synthesis of the scientific literature on the effects of disasters on dialysis populations. The indirect and direct effects of disasters on dialysis patients highlight the need for predisaster interventions at the patient and health care system levels. Particularly, educating patients about an emergency renal diet and offering early dialysis can help to mitigate the negative effects of disasters.

Time-varying Effects of GABRG1 and Maladaptive Peer Behavior on Externalizing Behavior from Childhood to Adulthood: Testing Gene × Environment × Development Effects.

Engagement in externalizing behavior is problematic. Deviant peer affiliation increases risk for externalizing behavior. Yet, peer effects vary across individuals and may differ across genes. This study determines gene × environment × development interactions as they apply to externalizing behavior from childhood to adulthood. A sample (n = 687; 68% male, 90% White) of youth from the Michigan Longitudinal Study was assessed from ages 10 to 25. Interactions between γ-amino butyric acid type A receptor γ1 subunit (GABRG1; rs7683876, rs13120165) and maladaptive peer behavior on externalizing behavior were examined using time-varying effect modeling. The findings indicate a sequential risk gradient in the influence of maladaptive peer behavior on externalizing behavior depending on the number of G alleles during childhood through adulthood. Individuals with the GG genotype are most vulnerable to maladaptive peer influences, which results in greater externalizing behavior during late childhood through early adulthood.

Alcoholic family marital heterogeneity aggregates different child behavior problems both pre- and postseparation.

Children of alcoholics (COAs) are at risk for elevated internalizing and externalizing symptoms. Yet, little is known about the familial and behavioral adjustments of COAs following parental separation. Using an ecological-transactional framework, we examined how multiple risk factors contributed to the formation of different alcoholic family structures and how living in heterogeneous family structures affected COAs' behavioral problems. The Michigan Longitudinal Study, a multiwave study on initially intact alcoholic and control families with preschool-age children (n = 503), was used to evaluate outcomes of offspring, when families either remained intact or were separated when the child was aged 12-14. Alcoholic families who later transitioned into stepfamilies were characterized with higher paternal antisociality, marital aggression, and serious family crises than alcoholic families that remained intact. COAs in stepfamilies (but not in single-parent families) exhibited higher levels of internalizing and externalizing symptoms in preadolescence compared with those in alcoholic intact families, in part because of elevated behavioral risk at age 3. Structural equation modeling indicated that the aggregated risk of stepfamily residence directly related to COAs' internalizing and indirectly related to COAs' externalizing problems, partially mediated by family stressors. Findings suggest targeting COAs in separated families for early intervention.

Childhood adversity, externalizing behavior, and substance use in adolescence: Mediating effects of anterior cingulate cortex activation during inhibitory errors.

Childhood adversity can negatively impact development across various domains, including physical and mental health. Adverse childhood experiences have been linked to aggression and substance use; however, developmental pathways to explain these associations are not well characterized. Understanding early precursors to later problem behavior and substance use can inform preventive interventions. The aim of the current study was to examine neurobiological pathways through which childhood adversity may lead to early adolescent problem behavior and substance use in late adolescence by testing two prospective models. Our first model found that early adolescent externalizing behavior mediates the association between childhood adversity and alcohol, cigarette, and marijuana use in late adolescence. Our second model found that activation in the anterior cingulate cortex (ACC) during an inhibitory control task mediates the association between childhood adversity and early adolescent externalizing behavior, with lower ACC activation associated with higher levels of adversity and more externalizing behavior. Together these findings indicate that the path to substance use in late adolescence from childhood adversity may operate through lower functioning in the ACC related to inhibitory control and externalizing behavior. Early life stressors should be considered an integral component in the etiology and prevention of early and problematic substance use.

Oxytocin Genotype Moderates the Impact of Social Support on Psychiatric Distress in Alcohol-Dependent Patients.

AIMS: The social environment strongly influences individual mental health. Individuals with strong social support systems tend to experience higher levels of well-being, lower levels of psychological distress and exhibit fewer psychiatric symptoms. However, there is a significant degree of individual variability as to the extent to which social support is beneficial to overall mental health. From a neurobiological perspective, it is suggested that the social hormone, oxytocin, may moderate the favorable effects of social interaction. To explore this possibility, we evaluated oxytocin genotype, social support and psychological health in a group of individuals diagnosed with DSM-IV alcohol dependence. METHODS: The associations between OXT genotype, social support and psychological health were analyzed in data from 269 adults diagnosed with DSM-IV alcohol dependence (25% female) admitted into residential treatment programs and outpatient centers in Warsaw, Poland. RESULTS: In line with past observations, we noted that psychiatric distress scores were negatively correlated with social support. Extending these observations, we uncovered a significant moderating effect of OXT genotype (rs2740210) on the relationship between social support and psychiatric distress. While G carriers displayed the predicted negative relationship between social support and psychiatric distress, T homozygotes failed to exhibit such a relationship. CONCLUSION: Genetically driven variation in oxytocin system functioning may influence the degree to which the beneficial effects of social support are felt in this population. These results have direct clinical relevance as enhancing social engagement to improve mental health may prove to be a less effective strategy in some patients owing to intrinsic factors. SHORT SUMMARY: The associations between oxytocin genotype, social support, and psychological health were analyzed in data from 269 adults diagnosed with DSM-IV alcohol dependence. A significant moderating effect of OXT genotype (rs2740210) on the relationship between social support and psychiatric distress was detected.

Pathways to Youth Behavior: The Role of Genetic, Neural, and Behavioral Markers.

Neural and temperamental mechanisms through which a genetic risk marker in the γ-amino butyric acid α2 receptor subunit (GABRA2) impacts adolescent functioning were investigated. Participants (N = 80; 29 female) completed an emotional word task during functional magnetic resonance imaging. Behavioral control, negative emotionality, and resiliency temperament constructs were assessed. Externalizing and internalizing problems were the outcomes. Those with the GABRA2 minor allele had reduced activation to positive words in the angular gyrus, middle temporal gyrus, and cerebellum, and to negative words in frontal, parietal, and occipital cortices. Reduced activation in the angular gyrus predicted greater negative emotionality and, in turn, elevated externalizing problems. Reduced activation in the inferior parietal cortex predicted greater resiliency and, in turn, low externalizing problems.

Brain Activity, Low Self-Control, and Delinquency: An fMRI Study of At-Risk Adolescents.

PURPOSE: A vast literature finds that low self-control is associated with a myriad of antisocial behaviors. Consequently, increasing attention has focused on the causes of low self-control. While criminologists have directed significant attention to studying its social causes, fewer studies have considered its neural bases. METHODS: We add to this nascent body of research by using data collected on an at-risk sample of adolescents participating in the ongoing Michigan Longitudinal Study. We examine the functioning of prefrontal and limbic regions of the brain during failed inhibitory control, assessed using the go/no-go task and functional magnetic resonance imaging, in relation to low self-control and self-reported delinquency. RESULTS: Results indicate that greater activation localized in the anterior cingulate cortex (ACC) during failed inhibitory control is negatively associated with low self-control. Moreover, the association between ACC activity and later delinquency is mediated through low self-control. CONCLUSIONS: Findings of this study demonstrate the utility of integrating neuroscientific and criminological perspectives on the causes of antisocial behavior. Concluding remarks address the theoretical and policy implications of the findings, as well as directions for future research.

A comprehensive framework for evaluating the environmental health and safety implications of engineered nanomaterials.

Engineered nanomaterials (ENM) are a growing aspect of the global economy, and their safe and sustainable development, use, and eventual disposal requires the capability to forecast and avoid potential problems. This review provides a framework to evaluate the health and safety implications of ENM releases into the environment, including purposeful releases such as for antimicrobial sprays or nano-enabled pesticides, and inadvertent releases as a consequence of other intended applications. Considerations encompass product life cycles, environmental media, exposed populations, and possible adverse outcomes. This framework is presented as a series of compartmental flow diagrams that serve as a basis to help derive future quantitative predictive models, guide research, and support development of tools for making risk-based decisions. After use, ENM are not expected to remain in their original form due to reactivity and/or propensity for hetero-agglomeration in environmental media. Therefore, emphasis is placed on characterizing ENM as they occur in environmental or biological matrices. In addition, predicting the activity of ENM in the environment is difficult due to the multiple dynamic interactions between the physical/chemical aspects of ENM and similarly complex environmental conditions. Others have proposed the use of simple predictive functional assays as an intermediate step to address the challenge of using physical/chemical properties to predict environmental fate and behavior of ENM. The nodes and interactions of the framework presented here reflect phase transitions that could be targets for development of such assays to estimate kinetic reaction rates and simplify model predictions. Application, refinement, and demonstration of this framework, along with an associated knowledgebase that includes targeted functional assay data, will allow better de novo predictions of potential exposures and adverse outcomes.

A time-varying effect model for examining group differences in trajectories of zero-inflated count outcomes with applications in substance abuse research.

This study proposes a time-varying effect model for examining group differences in trajectories of zero-inflated count outcomes. The motivating example demonstrates that this zero-inflated Poisson model allows investigators to study group differences in different aspects of substance use (e.g., the probability of abstinence and the quantity of alcohol use) simultaneously. The simulation study shows that the accuracy of estimation of trajectory functions improves as the sample size increases; the accuracy under equal group sizes is only higher when the sample size is small (100). In terms of the performance of the hypothesis testing, the type I error rates are close to their corresponding significance levels under all settings. Furthermore, the power increases as the alternative hypothesis deviates more from the null hypothesis, and the rate of this increasing trend is higher when the sample size is larger. Moreover, the hypothesis test for the group difference in the zero component tends to be less powerful than the test for the group difference in the Poisson component. Copyright © 2016 John Wiley & Sons, Ltd.

Beyond risk: Prospective effects of GABA Receptor Subunit Alpha-2 (GABRA2) × Positive Peer Involvement on adolescent behavior.

Research on Gene × Environment interactions typically focuses on maladaptive contexts and outcomes. However, the same genetic factors may also impact susceptibility to positive social contexts, leading to adaptive behavior. This paper examines whether the GABA receptor subunit alpha-2 (GABRA2) single nucleotide polymorphism rs279858 moderates the influence of positive peer affiliation on externalizing behavior and various forms of competence. Regions of significance were calculated to determine whether the form of the interaction supported differential susceptibility (increased sensitivity to both low and high positive peer affiliation) or vantage sensitivity (increased sensitivity to high positive peer affiliation). It was hypothesized that those carrying the homozygous minor allele (GG) would be more susceptible to peer effects. A sample (n = 300) of primarily male (69.7%) and White (93.0%) adolescents from the Michigan Longitudinal Study was assessed from ages 12 to 17. There was evidence for prospective Gene × Environment interactions in three of the four models. At low levels of positive peer involvement, those with the GG genotype were rated as having fewer adaptive outcomes, while at high levels they were rated as having greater adaptive outcomes. This supports differential susceptibility. Conceptualizing GABRA2 variants as purely risk factors may be inaccurate. Genetic differences in susceptibility to adaptive environmental exposures warrants further investigation.

Striatal dopaminergic reward response relates to age of first drunkenness and feedback response in at-risk youth.

Dopamine receptor concentrations, primarily in the striatum, are hypothesized to contribute to a developmental imbalance between subcortical and prefrontal control systems in emerging adulthood potentially biasing motivation and increasing risky behaviors. Positron emission tomography studies have found significant reductions in striatal dopamine D2 receptors, and blunted amphetamine-induced dopamine release, in substance users compared with healthy controls. Extant literature is limited and inconsistent concerning vulnerability associated with having a family history of substance abuse (FH+). Some studies have reported familial liability associated with higher dopamine receptor levels, reduced dopamine response to stimulant challenges and decreased response to oral alcohol. However, other reports have failed to find group differences based on family history. We explored the interaction of familial liability and behavioral risk with multi-modal molecular and neural imaging of the dopaminergic system. Forty-four young adult male subjects performed monetary incentive delay tasks during both [11 C]raclopride positron emission tomography and functional magnetic resonance imaging scans. FH+ subjects were identified as low (n = 24) or high risk (n = 9) based on early initiation of drunkenness. FH+ high-risk subjects exhibited heightened striatal dopamine response to monetary reward but did not differ in neural activations compared with FH+ low risk subjects and controls with no familial loading (n = 11). Across all subjects, a negative relationship was found between dopamine release and age of first drunkenness and a positive relationship with neural response to reward receipt. These results suggest that in at-risk individuals, higher dopamine transmission associated with monetary reward may represent a particularly useful neurobiological phenotype.

BOYS, EARLY RISK FACTORS FOR ALCOHOL PROBLEMS, AND THE DEVELOPMENT OF THE SELF: AN INTERCONNECTED MATRIX.

Alcohol-use disorders are a major public health issue worldwide. Although drinking and problematic alcohol use usually begins during adolescence, developmental origins of the disorder can be traced back to infancy and early childhood. Identification of early risk factors is essential to understanding developmental origins. Using data from the Michigan Longitudinal Study, an ongoing, prospective, high-risk family study, this article summarizes findings of family context and functioning of both children and parents. We draw attention to the development of the self, an understudied aspect of very young children being reared in alcoholic families that exacerbates exposure to high childhood adverse experiences. We also provide evidence demonstrating that young boys are embedded in a dynamic system of genes, epigenetic processes, brain organization, family dynamics, peers, community, and culture that strengthens risky developmental pathways if nothing is done to intervene during infancy and early childhood.

Characterization, detection, and counting of metal nanoparticles using flow cytometry.

There is a need to accurately detect, characterize, and quantify nanoparticles in suspensions. This study helps to understand the complex interactions between similar types of nanoparticles. Before initiating a study of metal nanoparticles, five submicron PS beads with sizes between 200 nm and 1 µm were used to derive a reference scale that was useful in evaluating the flow cytometer for functionality, sensitivity, resolution, and reproducibility. Side scatter intensity (SSC) from metal nanoparticles was obtained simultaneously from 405 nm and 488 nm lasers. The 405 nm laser generally yielded histogram distributions with smaller CVs, less side scatter intensity, better separation indices between beads and decreased scatter differences between different sized particles compared with the 488 nm laser. Submicron particles must be diluted to 10(6) and 10(7) particles/mL before flow cytometer analysis to avoid coincidence counting artifacts. When particles were too concentrated the following occurred: swarm, electronic overload, coincidence counting, activation of doublet discrimination and rejection circuitry, increase of mean SSC histogram distributions, alterations of SSC and pulse width histogram shape, decrease and fluctuations in counting rate and decrease or elimination of particulate water noise and 1 µm reference bead. To insure that the concentrations were in the proper counting range, the nanoparticle samples were mixed with a known concentration of 1 µm counting beads. Sequential dilutions of metal nanoparticles in a 1 µm counting bead suspension helped determine the diluted concentration needed for flow cytometer analysis. It was found that the original concentrated nanoparticle samples had to be diluted, between 1:10,000 and 1:100,000, before characterization by flow cytometry. The concentration of silver or gold nanoparticles in the undiluted sample were determined by comparing them with a known concentration (1.9 × 10(6) beads/mL) of 1 µm polystyrene reference beads.