Digital droplet assays-in which biological samples are compartmentalized into millions of femtoliter-volume droplets and interrogated individually-have generated enormous enthusiasm for their ability to detect biomarkers with single-molecule sensitivity. These assays have untapped potential for point-of-care diagnostics but are currently mainly confined to laboratory settings, due to the instrumentation necessary to serially generate, control, and measure tens of millions of droplets/compartments. To address this challenge, we developed an optofluidic platform that miniaturizes digital assays into a mobile format by parallelizing their operation. This technology is based on three key innovations: (i) the integration and parallel operation of a hundred droplet generators onto a single chip that operates >100× faster than a single droplet generator, (ii) the fluorescence detection of droplets at >100× faster than conventional in-flow detection using time domain-encoded mobile phone imaging, and (iii) the integration of on-chip delay lines and sample processing to allow serum-to-answer device operation. To demonstrate the power of this approach, we performed a duplex digital ELISA. We characterized the performance of this assay by first using spiked recombinant proteins in a complex media (FBS) and measured a limit of detection, 0.004 pg/mL (300 aM), a 1,000× improvement over standard ELISA and matching that of the existing laboratory-based gold standard digital ELISA system. We additionally measured endogenous GM-CSF and IL6 in human serum from n = 14 human subjects using our mobile duplex assay, and showed excellent agreement with the gold standard system ([Formula: see text]).

https://www.pnas.org/content/pnas/116/10/4489.full.pdf

Mobile platform for rapid sub-picogram-per-milliliter, multiplexed, digital droplet detection of proteins.

Background: A compelling number of studies, conducted in both children and adults, have reported an association between sleep disturbances/circadian sleep alterations and autism spectrum disorder (ASD); however, the data are sparse and the nature of this link is still unclear. The present review aimed to systematically collect the literature data relevant on sleep disturbances and circadian sleep dysrhythmicity related to ASD across all ages and to provide an integrative theoretical framework of their association. Methods: A systematic review of the MEDLINE, PubMed, and Cochrane databases was conducted from November 2018 to February 2019. The search strategies used were MeSH headings and keywords for "sleep-wake circadian rhythms" OR "circadian sleep disorders" OR "sleep-wake pattern" OR "sleep disorders" OR "melatonin" AND "autism spectrum disorder" OR "autism". Results: One hundred and three studies were identified, 15 regarded circadian sleep dysrhythmicity, 74 regarded sleep disturbances, and 17 regarded melatonin alterations in children and adults with ASD. Our findings suggested that autistic subjects frequently present sleep disturbances in particular short sleep duration, low sleep quality/efficiency, and circadian sleep desynchronization such as delayed phases and/or eveningness. Sleep disturbances and circadian sleep alterations have been related to the severity of autistic symptoms. Genetic studies have shown polymorphisms in circadian CLOCK genes and in genes involved in melatonin pathways in subjects with ASD. Conclusions: Sleep disturbances and circadian sleep alterations are frequent in subjects with autistic symptoms. These subjects have shown polymorphisms in clock genes expression and in genes involved in melatonin production. The impairment of circadian sleep regulation may increase the individual's vulnerability to develop symptoms of ASD by altering the sleep regulation in toto, which plays a key role in normal brain development. Even though controversies and "research gaps" are present in literature at this point, we may hypothesize a bidirectional relation between circadian sleep dysfunction and ASD. In particular, circadian sleep dysrhythmicity may predispose to develop ASD symptoms and vice versa within a self-reinforcing feedback loop. By targeting sleep disturbances and circadian sleep dysrhythmicity, we may improve treatment strategies for both children and adults with ASD.

/pdf/systematic-review-of-sleep-disturbances-and-circadian-sleep-17zcbp60wh.pdf

Systematic Review of Sleep Disturbances and Circadian Sleep Desynchronization in Autism Spectrum Disorder: Toward an Integrative Model of a Self-Reinforcing Loop

Tuberculosis (TS) is now the worlds foremost cause of death from a single infectious agent with nearly 90 million new cases and 30 million deaths annually. The economic impact of disease by way of direct and indirect costs of contact investigations, costs of TB screening and preventive therapy programs, costs of hospital and institutional infection control programs and costs to patients in lost income, is enormous. An increasing number of cases converting to multidrug resistant tuberculosis could escalate these costs dramatically. [1]

The magnitude of the problem.

Pediatric sleep health: It matters, and so does how we define it.

Sleep disturbances in children with autism spectrum disorder (ASD) are significantly more prevalent than found in typically developing (TD) children. Given the detrimental impact of poor sleep on cognitive, emotional, and behavioral functioning, it is imperative to screen and assess for sleep disturbances in this population. In this paper, we describe the screening and assessment process, as well as specific measures commonly used for assessing sleep in children with ASD. Advantages and limitations for use in children with ASD are discussed. While subjective measures, such as parent-report questionnaires and sleep diaries, are the most widely used, more objective measures such as actigraphy, polysomnography, and videosomnography provide additional valuable information for both diagnostic purposes and treatment planning. These objective measures, nonetheless, are limited by cost, availability, and feasibility of use with children with ASD. The current review provides an argument for the complementary uses of both subjective and objective measures of sleep specifically for use in children with ASD.

https://www.mdpi.com/2227-9067/4/8/72/pdf

Assessment of Sleep in Children with Autism Spectrum Disorder.

We studied 28 adolescents/young adults with autism spectrum disorders (ASD) and 13 age/sex matched individuals of typical development (TD). Structured sleep histories, validated questionnaires, actigraphy (4 weeks), and salivary cortisol and melatonin (4 days each) were collected. Compared to those with TD, adolescents/young adults with ASD had longer sleep latencies and more difficulty going to bed and falling asleep. Morning cortisol, evening cortisol, and the morning-evening difference in cortisol did not differ by diagnosis (ASD vs. TD). Dim light melatonin onsets (DLMOs) averaged across participants were not different for the ASD and TD participants. Average participant scores indicated aspects of poor sleep hygiene in both groups. Insomnia in ASD is multifactorial and not solely related to physiological factors.

/pdf/characterizing-sleep-in-adolescents-and-adults-with-autism-j0xg7tahvi.pdf

Characterizing Sleep in Adolescents and Adults with Autism Spectrum Disorders.

Aim: Digital ELISA-based assays for blood biomarkers of neurological disease are on the verge of clinical use. Here, we aimed to determine whether different preanalytical blood processing technique...

Neuro biomarker levels measured with high-sensitivity digital ELISA differ between serum and plasma.

Teaching children with autism spectrum disorder how to sleep better: A pilot educational program for parents.

Background: Historically, limited sensitivity associated with traditional immunoassay methods has prevented the use of brain-specific proteins as blood biomarkers of traumatic brain injury (TBI) during triage, as these proteins exhibit low circulating concentrations. Digital ELISA is a newly-developed technique that is up to 1000 times more sensitive than conventional ELISA methods. The purpose of this study was to determine whether the use of digital ELISA over conventional ELISA improves the performance of brain-specific proteins as blood biomarkers of TBI during triage.Methods: Blood was sampled from TBI patients (n = 13) at emergency department admission, as well as from neurologically normal controls (n = 72). Serum levels of two brain-specific proteins, neurofilament light chain (NfL) and Tau, were measured via digital ELISA. Estimated conventional ELISA measures were generated by adjusting values according to the lower limits of detection achievable with commercially available conventional ELISA assays, and receiver operating characteristic (ROC) analysis was used to compare the diagnostic performance of digital ELISA measures to estimated conventional ELISA measures in terms of their ability to discriminate between TBI patients and controls.Results: Used in combination, digital ELISA measures of NfL and Tau could discriminate between groups with 100% sensitivity and 91.7% specificity. Estimated conventional ELISA measures could only discriminate between groups with 7.7% sensitivity and 94.4% specificity. This difference in diagnostic performance was statistically significant when comparing areas under ROC curves.Conclusions: The use of digital ELISA over conventional ELISA methods improves the diagnostic performance of circulating brain-specific proteins for detection of TBI during triage.

Use of high-sensitivity digital ELISA improves the diagnostic performance of circulating brain-specific proteins for detection of traumatic brain injury during triage

Megan L. Alder

Papers

Characterizing Sleep in Adolescents and Adults with Autism Spectrum Disorders.

Neuro biomarker levels measured with high-sensitivity digital ELISA differ between serum and plasma.

Teaching children with autism spectrum disorder how to sleep better: A pilot educational program for parents.

Use of high-sensitivity digital ELISA improves the diagnostic performance of circulating brain-specific proteins for detection of traumatic brain injury during triage

Diagnosis of ischemic stroke using circulating levels of brain-specific proteins measured via high-sensitivity digital ELISA.