The impact of family physicians in rural maternity care.

BACKGROUND: Reduced access to maternity care in rural areas of the United States presents a significant burden to pregnant persons and infants. The objective of this study was to estimate the impact of family physicians (FPs) on access to maternity care in rural United States hospitals, especially where other providers may not be available. METHODS: We administered a survey to 216 rural hospitals in 10 US states inquiring about the number of babies delivered from 2013 to 2017, the types of delivering physicians, and the maternity services offered. We calculated the percentage of rural hospitals in our sample where FPs performed vaginal deliveries, cesareans, and vaginal births after cesarean (VBACs), and the percentage of all babies delivered by FPs. We estimated the distance patients would have to travel for care if FPs were not providing care locally. RESULTS: The final study population consisted of 185 rural hospitals. FPs delivered babies in 67% of these hospitals and were the only physicians who delivered babies in 27% of these hospitals. FPs provided VBAC at 18% and cesarean birth services at 46% of the rural hospitals, but with wide geographic differences. Many patients would have to drive an average of 86 miles round-trip to access care if those FPs were to stop delivering. CONCLUSIONS: Family physicians are essential providers of maternity care in the rural United States. Family Medicine residency programs should ensure that trainees who intend to practice in rural locations have adequate maternity care training to maintain and expand access to maternity care for rural patients and their families.

A heart-team hybrid approach for atrial fibrillation: a single-centre long-term clinical outcome cohort study.

OBJECTIVES: The Cox-maze IV is the gold standard for surgical ablation of atrial fibrillation (AF). A heart-team hybrid approach using selected epicardial thoracoscopic surgical ablations and completion endocardial ablations to replicate the Cox-maze IV lesion set has gained popularity and early results have been promising. We herein report our single-centre long-term clinical outcomes using the heart-team hybrid approach with 455 patients. METHODS: From 1 March 2013 to 1 July 2019, we prospectively collected data on all patients referred to our heart team for rhythm-control strategy for AF. Baseline characteristics, procedural complications and long-term freedom from AF (FFAF) both on and off anti-arrhythmic drug therapy were analysed. Ambulatory monitoring (>7 days) was obtained at 3 months and annually thereafter. RESULTS: Four hundred and fifty-five patients completed the hybrid approach. Four hundred and forty-five (97.8%) patients had non-paroxysmal AF (long-standing persistent AF n = 249, 54.7%; persistent AF n = 196, 43.1%; paroxysmal AF n = 10, 2.2%). Average duration of AF was 5.9 ± 6.1 years. Average left atrial diameter was 4.8 ± 0.8 cm. FFAF at 3, 12, 24 and 36 months was 92%, 87%, 81% and 72%, respectively. FFAF without the use of anti-arrhythmic medications was 75%, 81%, 76% and 66%. Any surgical complications occurred in 28 (6.1%) patients. CONCLUSIONS: A heart-team hybrid strategy for the treatment of AF is safe and effective. In a predominantly non-paroxysmal population with AF, at the 3-year follow-up, FFAF in patients on and off anti-arrhythmic drugs approaches that of patients who had the Cox-maze IV.

Prospective Evaluation of Adverse Event Recognition Systems in Twitter: Results from the Web-RADR Project.

INTRODUCTION: A large number of studies on systems to detect and sometimes normalize adverse events (AEs) in social media have been published, but evidence of their practical utility is scarce. This raises the question of the transferability of such systems to new settings. OBJECTIVES: The aims of this study were to develop an AE recognition system, prospectively evaluate its performance on an external benchmark dataset and identify potential factors influencing the transferability of AE recognition systems. METHODS: A pipeline based on dictionary lookups and logistic regression classifiers was developed using a proprietary dataset of 196,533 Tweets manually annotated for AE relations and prospectively evaluated the system on the publicly available WEB-RADR reference dataset, exploring different aspects affecting transferability. RESULTS: Our system achieved 0.53 precision, 0.52 recall and 0.52 F1-score on the development test set; however, when applied to the WEB-RADR reference dataset, system performance dropped to 0.38 precision, 0.20 recall and 0.26 F1-score. Similarly, a previously published method aiming at automatically detecting adverse event posts reported 0.5 precision, 0.92 recall and 0.65 F1-score on thus another dataset, while performance on the WEB-RADR reference dataset was reduced to 0.37 precision, 0.63 recall and 0.46 F1-score. We identified four potential factors leading to poor transferability: overfitting, selection bias, label bias and prevalence. CONCLUSION: We warn the community about a potentially large discrepancy between the expected performance of automated AE recognition systems based on published results and the actual observed performance on independent data. This study highlights the difficulty of implementing an all-purpose system for automatic adverse event recognition in Twitter, which could explain the lack of such systems in practical pharmacovigilance settings. Our recommendation is to use benchmark independent datasets, such as the WEB-RADR reference, to investigate the transferability of the adverse event recognition systems and ultimately enforce rigorous comparisons across studies on the task.

Correction: Social Media Surveillance of Multiple Sclerosis Medications Used During Pregnancy and Breastfeeding: Content Analysis.

[This corrects the article DOI: .].

Social Media Surveillance of Multiple Sclerosis Medications Used During Pregnancy and Breastfeeding: Content Analysis.

BACKGROUND: Multiple sclerosis (MS) is a chronic neurological disease occurring mostly in women of childbearing age. Pregnant women with MS are usually excluded from clinical trials; as users of the internet, however, they are actively engaged in threads and forums on social media. Social media provides the potential to explore real-world patient experiences and concerns about the use of medicinal products during pregnancy and breastfeeding. OBJECTIVE: This study aimed to analyze the content of posts concerning pregnancy and use of medicines in online forums; thus, the study aimed to gain a thorough understanding of patients' experiences with MS medication. METHODS: Using the names of medicinal products as search terms, we collected posts from 21 publicly available pregnancy forums, which were accessed between March 2015 and March 2018. After the identification of relevant posts, we analyzed the content of each post using a content analysis technique and categorized the main topics that users discussed most frequently. RESULTS: We identified 6 main topics in 70 social media posts. These topics were as follows: (1) expressing personal experiences with MS medication use during the reproductive period (55/70, 80%), (2) seeking and sharing advice about the use of medicines (52/70, 74%), (3) progression of MS during and after pregnancy (35/70, 50%), (4) discussing concerns about MS medications during the reproductive period (35/70, 50%), (5) querying the possibility of breastfeeding while taking MS medications (30/70, 42%), and (6) commenting on communications with physicians (26/70, 37%). CONCLUSIONS: Overall, many pregnant women or women considering pregnancy shared profound uncertainties and specific concerns about taking medicines during the reproductive period. There is a significant need to provide advice and guidance to MS patients concerning the use of medicines in pregnancy and postpartum as well as during breastfeeding. Advice must be tailored to the circumstances of each patient and, of course, to the individual medicine. Information must be provided by a trusted source with relevant expertise and made publicly available.

Recommendations for the Use of Social Media in Pharmacovigilance: Lessons from IMI WEB-RADR.

Over a period of 3 years, the European Union's Innovative Medicines Initiative WEB-RADR project has explored the value of social media (i.e., information exchanged through the internet, typically via online social networks) for identifying adverse events as well as for safety signal detection. Many patients and clinicians have taken to social media to discuss their positive and negative experiences of medications, creating a source of publicly available information that has the potential to provide insights into medicinal product safety concerns. The WEB-RADR project has developed a collaborative English language workspace for visualising and analysing social media data for a number of medicinal products. Further, novel text and data mining methods for social media analysis have been developed and evaluated. From this original research, several recommendations are presented with supporting rationale and consideration of the limitations. Recommendations for further research that extend beyond the scope of the current project are also presented.

Recommendations on the Use of Mobile Applications for the Collection and Communication of Pharmaceutical Product Safety Information: Lessons from IMI WEB-RADR.

Over a period of 3 years, the European Union's Innovative Medicines Initiative WEB-RADR (Recognising Adverse Drug Reactions; https://web-radr.eu/ ) project explored the value of two digital tools for pharmacovigilance (PV): mobile applications (apps) for reporting the adverse effects of drugs and social media data for its contribution to safety signalling. The ultimate intent of WEB-RADR was to provide policy, technical and ethical recommendations on how to develop and implement such digital tools to enhance patient safety. Recommendations relating to the use of mobile apps for PV are summarised in this paper. There is a presumption amongst at least some patients and healthcare professionals that information ought to be accessed and reported from any setting, including mobile apps. WEB-RADR has focused on the use of such technology for reporting suspected adverse drug reactions and for broadcasting safety information to its users, i.e. two-way risk communication. Three apps were developed and publicly launched within Europe as part of the WEB-RADR project and subsequently assessed by a range of stakeholders to determine their value as effective tools for improving patient safety; a fourth generic app was later piloted in two African countries. The recommendations from the development and evaluation of the European apps are presented here with supporting considerations, rationales and caveats as well as suggested areas for further research.

Accurate and selective quantification of anthrax protective antigen in plasma by immunocapture and isotope dilution mass spectrometry.

Anthrax protective antigen (83 kDa, PA83) is an essential component of two major binary toxins produced by Bacillus anthracis, lethal toxin (LTx) and edema toxin (ETx). During infection, LTx and ETx contribute to immune collapse, endothelial dysfunction, hemorrhage and high mortality. Following protease cleavage on cell receptors or in circulation, the 20 kDa (PA20) N-terminus is released, activating the 63 kDa (PA63) form which binds lethal factor (LF) and edema factor (EF), facilitating their entry into their cellular targets. Several ELISA-based PA methods previously developed are primarily qualitative or semi-quantitative. Here, we combined protein immunocapture, tryptic digestion and isotope dilution liquid chromatography-mass spectrometry (LC-MS/MS), to develop a highly selective and sensitive method for detection and accurate quantification of total-PA (PA83 + PA63) and PA83. Two tryptic peptides in the 63 kDa region measure total-PA and three in the 20 kDa region measure PA83 alone. Detection limits range from 1.3-2.9 ng mL-1 PA in 100 µL of plasma. Spiked recovery experiments with combinations of PA83, PA63, LF and EF in plasma showed that PA63 and PA83 were quantified accurately against the PA83 standard and that LF and EF did not interfere with accuracy. Applied to a study of inhalation anthrax in rhesus macaques, total-PA suggested triphasic kinetics, similar to that previously observed for LF and EF. This study is the first to report circulating PA83 in inhalation anthrax, typically at less than 4% of the levels of PA63, providing the first evidence that activated PA63 is the primary form of PA throughout infection.

Enabling social listening for cardiac safety monitoring: Proceedings from a drug information association-cardiac safety research consortium cosponsored think tank.

This white paper provides a summary of the presentations and discussions from a think tank on "Enabling Social Listening for Cardiac Safety Monitoring" trials that was cosponsored by the Drug Information Association and the Cardiac Safety Research Consortium, and held at the White Oak headquarters of the US Food and Drug Administration on June 3, 2016. The meeting's goals were to explore current methods of collecting and evaluating social listening data and to consider their applicability to cardiac safety surveillance. Social listening is defined as the act of monitoring public postings on the Internet. It has several theoretical advantages for drug and device safety. First, these include the ability to detect adverse events that are "missed" by traditional sources and the ability to detect adverse events sooner than would be allowed by traditional sources, both by affording near-real-time access to data from culturally and geographically diverse sources. Social listening can also potentially introduce a novel patient voice into the conversation about drug safety, which could uniquely augment understanding of real-world medication use obtained from more traditional methodologies. Finally, it can allow for access to information about drug misuse and diversion. To date, the latter 2 of these have been realized. Although regulators from the Food and Drug Administration and the United Kingdom's Medicines and Healthcare Products Regulatory Agency participated in the think tank along with representatives from industry, academia, and patient groups, this article should not be construed to constitute regulatory guidance.

Immunocapture isotope dilution mass spectrometry in response to a pandemic influenza threat.

As a result of recent advances in mass spectrometry-based protein quantitation methods, these techniques are now poised to play a critical role in rapid formulation of pandemic influenza vaccines. Analytical techniques that have been developed and validated on seasonal influenza strains can be used to increase the quality and decrease the time required to deliver protective pandemic vaccines to the global population. The emergence of a potentially pandemic avian influenza A (H7N9) virus in March of 2013, prompted the US public health authorities and the vaccine industry to initiate production of a pre-pandemic vaccine for preparedness purposes. To this end, we evaluated the feasibility of using immunocapture isotope dilution mass spectrometry (IC-IDMS) to evaluate the suitability of the underlying monoclonal and polyclonal antibodies (mAbs and pAbs) for their capacity to isolate the H7 hemagglutinin (HA) in this new vaccine for quantification by IDMS. A broad range of H7 capture efficiencies was observed among mAbs tested by IC-IDMS with FR-545, 46/6, and G3 A533 exhibiting the highest cross-reactivity capabilities to H7 of A/Shanghai/2/2013. MAb FR-545 was selected for continued assessment, evaluated by IC-IDMS for mAb reactivity against H7 in the H7N9 candidate vaccine virus and compared with/to reactivity to the reference polyclonal antiserum in allantoic fluid, purified whole virus, lyophilized whole virus and final detergent-split monovalent vaccine preparations for vaccine development. IC-IDMS assessment of FR-545 alongside IC-IDMS using the reference polyclonal antiserum to A/Shanghai/2/2013 and with the regulatory SRID method showed strong correlation and mAb IC-IDMS could have played an important role in the event a potential surrogate potency test was required to be rapidly implemented.

Quantification of Influenza Neuraminidase Activity by Ultra-High Performance Liquid Chromatography and Isotope Dilution Mass Spectrometry.

Mounting evidence suggests that neuraminidase's functionality extends beyond its classical role in influenza virus infection and that antineuraminidase antibodies offer protective immunity. Therefore, a renewed interest in the development of neuraminidase (NA)-specific methods to characterize the glycoprotein and evaluate potential advantages for NA standardization in influenza vaccines has emerged. NA displays sialidase activity by cleaving off the terminal N-acetylneuraminic acid on α-2,3 or α-2,6 sialic acid containing receptors of host cells. The type and distribution of these sialic acid containing receptors is considered to be an important factor in transmission efficiency of influenza viruses between and among host species. Changes in hemagglutinin (HA) binding and NA specificity in reassortant viruses may be related to the emergence of new and potentially dangerous strains of influenza. Current methods to investigate neuraminidase activity use small derivatized sugars that are poor models for natural glycoprotein receptors and do not provide information on the linkage specificity. Here, a novel approach for rapid and accurate quantification of influenza neuraminidase activity is achieved utilizing ultra-high performance liquid chromatography (UPLC) and isotope dilution mass spectrometry (IDMS). Direct LC-MS/MS quantification of NA-released sialic acid provides precise measurement of influenza neuraminidase activity over a range of substrates. The method provides exceptional sensitivity and specificity with a limit of detection of 0.38 µM for sialic acid and the capacity to obtain accurate measurements of specific enzyme activity preference toward α-2,3-sialyllactose linkages, α-2,6-sialyllactose linkages, or whole glycosylated proteins such as fetuin.

Evaluation of Facebook and Twitter Monitoring to Detect Safety Signals for Medical Products: An Analysis of Recent FDA Safety Alerts.

INTRODUCTION: The rapid expansion of the Internet and computing power in recent years has opened up the possibility of using social media for pharmacovigilance. While this general concept has been proposed by many, central questions remain as to whether social media can provide earlier warnings for rare and serious events than traditional signal detection from spontaneous report data. OBJECTIVE: Our objective was to examine whether specific product-adverse event pairs were reported via social media before being reported to the US FDA Adverse Event Reporting System (FAERS). METHODS: A retrospective analysis of public Facebook and Twitter data was conducted for 10 recent FDA postmarketing safety signals at the drug-event pair level with six negative controls. Social media data corresponding to two years prior to signal detection of each product-event pair were compiled. Automated classifiers were used to identify each 'post with resemblance to an adverse event' (Proto-AE), among English language posts. A custom dictionary was used to translate Internet vernacular into Medical Dictionary for Regulatory Activities (MedDRA®) Preferred Terms. Drug safety physicians conducted a manual review to determine causality using World Health Organization-Uppsala Monitoring Centre (WHO-UMC) assessment criteria. Cases were also compared with those reported in FAERS. FINDINGS: A total of 935,246 posts were harvested from Facebook and Twitter, from March 2009 through October 2014. The automated classifier identified 98,252 Proto-AEs. Of these, 13 posts were selected for causality assessment of product-event pairs. Clinical assessment revealed that posts had sufficient information to warrant further investigation for two possible product-event associations: dronedarone-vasculitis and Banana Boat Sunscreen--skin burns. No product-event associations were found among the negative controls. In one of the positive cases, the first report occurred in social media prior to signal detection from FAERS, whereas the other case occurred first in FAERS. CONCLUSIONS: An efficient semi-automated approach to social media monitoring may provide earlier insights into certain adverse events. More work is needed to elaborate additional uses for social media data in pharmacovigilance and to determine how they can be applied by regulatory agencies.

Social Media Listening for Routine Post-Marketing Safety Surveillance.

INTRODUCTION: Post-marketing safety surveillance primarily relies on data from spontaneous adverse event reports, medical literature, and observational databases. Limitations of these data sources include potential under-reporting, lack of geographic diversity, and time lag between event occurrence and discovery. There is growing interest in exploring the use of social media ('social listening') to supplement established approaches for pharmacovigilance. Although social listening is commonly used for commercial purposes, there are only anecdotal reports of its use in pharmacovigilance. Health information posted online by patients is often publicly available, representing an untapped source of post-marketing safety data that could supplement data from existing sources. OBJECTIVES: The objective of this paper is to describe one methodology that could help unlock the potential of social media for safety surveillance. METHODS: A third-party vendor acquired 24 months of publicly available Facebook and Twitter data, then processed the data by standardizing drug names and vernacular symptoms, removing duplicates and noise, masking personally identifiable information, and adding supplemental data to facilitate the review process. The resulting dataset was analyzed for safety and benefit information. RESULTS: In Twitter, a total of 6,441,679 Medical Dictionary for Regulatory Activities (MedDRA(®)) Preferred Terms (PTs) representing 702 individual PTs were discussed in the same post as a drug compared with 15,650,108 total PTs representing 946 individual PTs in Facebook. Further analysis revealed that 26 % of posts also contained benefit information. CONCLUSION: Social media listening is an important tool to augment post-marketing safety surveillance. Much work remains to determine best practices for using this rapidly evolving data source.

Increasing Patient Engagement in Pharmacovigilance Through Online Community Outreach and Mobile Reporting Applications: An Analysis of Adverse Event Reporting for the Essure Device in the US.

BACKGROUND: Preparing and submitting a voluntary adverse event (AE) report to the US Food and Drug Administration (FDA) for a medical device typically takes 40 min. User-friendly Web and mobile reporting apps may increase efficiency. Further, coupled with strategies for direct patient involvement, patient engagement in AE reporting may be improved. In 2012, the FDA Center for Devices and Radiologic Health (CDRH) launched a free, public mobile AE reporting app, MedWatcher, for patients and clinicians. During the same year, a patient community on Facebook adopted the app to submit reports involving a hysteroscopic sterilization device, brand name Essure®. METHODS: Patient community outreach was conducted to administrators of the group "Essure Problems" (approximately 18,000 members as of June 2015) to gather individual case safety reports (ICSRs). After agreeing on key reporting principles, group administrators encouraged members to report via the app. Semi-structured forms in the app mirrored fields of the MedWatch 3500 form. ICSRs were transmitted to CDRH via an electronic gateway, and anonymized versions were posted in the app. Data collected from May 11, 2013 to December 7, 2014 were analyzed. Narrative texts were coded by trained and certified MedDRA coders (version 17). Descriptive statistics and metrics, including VigiGrade completeness scores, were analyzed. Various incentives and motivations to report in the Facebook group were observed. RESULTS: The average Essure AE report took 11.4 min (±10) to complete. Submissions from 1349 women, average age 34 years, were analyzed. Serious events, including hospitalization, disability, and permanent damage after implantation, were reported by 1047 women (77.6 %). A total of 13,135 product-event pairs were reported, comprising 327 unique preferred terms, most frequently fatigue (n = 491), back pain (468), and pelvic pain (459). Important medical events (IMEs), most frequently mental impairment (142), device dislocation (108), and salpingectomy (62), were reported by 598 women (44.3 %). Other events of interest included loss of libido (n = 115); allergy to metals (109), primarily nickel; and alopecia (252). VigiGrade completeness scores were high, averaging 0.80 (±0.15). Reports received via the mobile app were considered "well documented" 55.9 % of the time, compared with an international average of 13 % for all medical products. On average, there were 15 times more reports submitted per month via the app with patient community support versus traditional pharmacovigilance portals. CONCLUSIONS: Outreach via an online patient community, coupled with an easy-to-use app, allowed for rapid and detailed ICSRs to be submitted, with gains in efficiency. Two-way communication and public posting of narratives led to successful engagement within a Motivation-Incentive-Activation-Behavior framework, a conceptual model for successful crowdsourcing. Reports submitted by patients were considerably more complete than those submitted by physicians in routine spontaneous reports. Further research is needed to understand how biases operate differently from those of traditional pharmacovigilance.

Simultaneous Identification and Susceptibility Determination to Multiple Antibiotics of Staphylococcus aureus by Bacteriophage Amplification Detection Combined with Mass Spectrometry.

The continued advance of antibiotic resistance in clinically relevant bacterial strains necessitates the development and refinement of assays that can rapidly and cost-effectively identify bacteria and determine their susceptibility to a panel of antibiotics. A methodology is described herein that exploits the specificity and physiology of the Staphylococci bacteriophage K to identify Staphylococcus aureus (S. aureus) and determine its susceptibility to clindamycin and cefoxitin. The method uses liquid chromatography-mass spectrometry to monitor the replication of bacteriophage after it is used to infect samples thought to contain S. aureus. Amplification of bacteriophage K indicates the sample contains S. aureus, for it is only in the presence of a suitable host that bacteriophage K can amplify. If bacteriophage amplification is detected in samples containing the antibiotics clindamycin or cefoxitin, the sample is deemed to be resistant to these antibiotics, respectively, for bacteriophage can only amplify in a viable host. Thus, with a single work flow, S. aureus can be detected in an unknown sample and susceptibility to clindamycin and cefoxitin can be ascertained. This Article discusses implications for the use of bacteriophage amplification in the clinical laboratory.

Viable Staphylococcus aureus quantitation using ¹5N metabolically labeled bacteriophage amplification coupled with a multiple reaction monitoring proteomic workflow.

A multiple reaction monitoring liquid chromatography method with tandem mass spectrometric detection for quantitation of Staphylococcus aureus via phage amplification detection is described. This phage amplification detection method enables rapid and accurate quantitation of viable S. aureus by detecting an amplified capsid protein from a specific phage. A known amount of metabolically labeled (15)N reference bacteriophage, utilized as the input phage and as the internal standard for quantitation, was spiked into S. aureus samples. Following a 2-h incubation, the sample was subjected to a 3-min rapid trypsin digest and analyzed by high-throughput liquid chromatography tandem mass spectrometric detection targeting peptides unique to both the (15)N (input phage) and (14)N (progeny phage) capsid proteins. Quantitation was achieved by comparing peak areas of target peptides from the metabolically labeled (15)N bacteriophage peptide internal standard with that of the wild-type (14)N peptides that were produced by phage amplification and subsequent digestion when the host bacteria was present. This approach is based on the fact that a labeled species differs from the unlabeled one in terms of its mass but exhibits almost identical chemical properties such as ion yields and retention times. A 6-point calibration curve for S. aureus concentration was constructed with standards ranging from 5.0 × 10(4) colony forming units (CFU) ml(-1) to 2.0 × 10(6) CFU ml(-1), with the (15)N reference phage spiked at a concentration of 1.0 × 10(9) plaque forming units (PFU) ml(-1). Amplification with (15)N bacteriophage coupled with LC-MS/MS detection offers speed (3 h total analysis time), sensitivity (LOD: < 5.0 × 10(4) CFU ml(-1)), accuracy, and precision for quantitation of S. aureus.

Quantification of immunoreactive viral influenza proteins by immunoaffinity capture and isotope-dilution liquid chromatography-tandem mass spectrometry.

An immunocapture isotope dilution mass spectrometry (IC-IDMS) method was developed to quantify antibody-bound influenza hemagglutinins (HA) in trivalent influenza vaccines (TIV). Currently, regulatory potency requirements for TIV require HA quantification based on the single radial immunodiffusion (SRID) assay, which is time-consuming, laborious, and requires production of large quantities of reagents globally. In IC-IDMS, antiserum to the HA of interest captured viral proteins that were in the correct conformation to be recognized by the antibodies. The captured proteins were digested, and evolutionarily conserved tryptic peptides were quantified using isotope-dilution liquid chromatography-tandem mass spectrometry. IC-IDMS relies on antibody-antigen binding similar to SRID but incorporates the accuracy and precision of IDMS. Polyclonal antibodies (pAb-H3) prepared by injection of sheep with purified H3 HA captured 82.9% (55.26 fmol/µL) of the total H3 HA (66.69 fmol/µL) from the commercial TIV and 93.6% (57.23 fmol/µL) of the total H3 HA (61.14 fmol/µL) in purified virus. While other HA (H1, B), neuraminidase (N1, N2, NB), viral matrix proteins, and nucleoproteins were also captured by this antiserum, our results were not affected due to the specificity of the mass spectrometer. IC-IDMS is an accurate, precise, sensitive, and selective method to measure antibody-bound HA in purified virus and commercial vaccines.

Detection of Staphylococcus aureus using 15N-labeled bacteriophage amplification coupled with matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry.

A novel approach to rapid bacterial detection using an isotopically labeled (15)N bacteriophage and matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS) is introduced. Current phage amplification detection (PAD) via mass spectrometric analysis is limited because host bacteria must be inoculated with low phage titers in such a way that initial infecting phage concentrations must be below the detection limit of the instrument, thus lengthening incubation times. Additionally, PAD techniques cannot distinguish inoculate input phage from output phage which can increase the possibility of false positive results. Here, we report a rapid and accurate PAD approach for identification of Staphylococcus aureus via detection of bacteriophage capsid proteins. This approach uses both a wild-type (14)N and a (15)N-isotopically labeled S. aureus-specific bacteriophage. High (15)N phage titers, above our instrument's detection limits, were used to inoculate S. aureus. MALDI-TOF MS detection of the (14)N progeny capsid proteins in the phage-amplified culture indicated the presence of the host bacteria. Successful phage amplification was observed after 90 min of incubation. The amplification was observed by both MALDI-TOF MS analysis and by standard plaque assay measurements. This method overcomes current limitations by improving analysis times while increasing selectivity when compared to previously reported PAD methodologies.