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We present a protocol to generate highly multiplexed spatial data at cellular and subcellular resolutions using iterative indirect immunofluorescence imaging (4i). We describe streamlined steps for using 4i across fixed cultured cells, formalin-fixed paraffin-embedded (FFPE) tissue sections, and metaphase chromosome spreads. We detail procedures for sample preparation, antibody and DNA staining, immunofluorescence imaging, antibody elution, and image processing. This protocol is adapted for high-throughput analysis of fixed cultured cells and addresses sample-specific challenges such as intrinsic tissue autofluorescence and chromosome fragility. For complete details on the use and execution of this protocol for fixed cultured cells, please refer to Comandante-Lou et al.1.
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New Jersey (NJ) Safe Schools Program (NJSS) provides code-required trainings for NJ teachers supervising students in work-based learning (WBL) experiences. During the COVID-19 pandemic, increased use of chemical cleaning, sanitization, and disinfectant products (CSDPs) led to ventilation and other health concerns. NJSS conducted two surveys of newer NJ WBL teachers between October 2021 and June 2023, with a follow-up in fall 2022 for those who completed initial surveys before summer 2022. This study focused on questions regarding workspace ventilation; respirator and CSDPs trainings; awareness of government resources for safety and health (S&H); and demographics. Over 65% of participant classrooms have operable windows or ventilation systems, while 13% of school salons have ceiling fans. Half of participants didn't receive training on handling CSDPs, although 90% were aware of government S&H resources. Data suggested teachers have well-ventilated workspaces and are aware of different resources available, which is important to continue post COVID-19.
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During the COVID-19 pandemic, there was an increased reported use of chemical cleaning, sanitizing, and disinfecting products (CSDPs), which created public concerns about negative health consequences for both children and adults in public schools. A subset of newer teachers shared experiences regarding safety and health (S&H) while working in school-based settings through a series of online surveys. Surveys were provided to teachers who completed work-based learning supervisory trainings provided by the New Jersey Safe Schools Program between October 2021 and June 2023. The participants answered questions focusing on CSDPs purchased for school use, their attitudes towards CSDPs, their use of personal protective equipment, and symptoms employees may have had due to CSDPs. A total of 205 teacher participants successfully completed the surveys. Over 25% of the teachers did not know where their CSDPs originated from, as they were provided by the school. Most participants "sometimes", "not often", or "never" read labels for CSDP ingredients or looked them up on healthy product apps. The participants (60%) tended to wear gloves while cleaning/disinfecting but did not wear masks. A third of the participants experienced respiratory health problems after working at school. Overall, the data suggest that more education on S&H regarding CSDPs needs to be provided to New Jersey teachers.
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Pandemias , Maestros , Niño , Adulto , Humanos , New Jersey , Instituciones Académicas , ActitudRESUMEN
BACKGROUND: Vaccine hesitancy remains a societal problem, including during the COVID-19 pandemic. New Jersey (NJ) Safe Schools Program provides work-based learning training to supervisory-level career-technical-vocational education teachers and administrators who have to consider varied state and local mandates concerning COVID-19 vaccination and exemptions. METHODS: In early 2022, we distributed an online survey via PsychData to individuals trained between 2014 and 2022 to understand NJ teachers' practices and concerns regarding COVID-19 vaccines. Overall, 269 completed the survey. We stratified data by vaccination status, number of doses, booster status, age, teaching experience, gender, race, county of work, and COVID-19 diagnosis status. RESULTS: Overall, results suggested differences in COVID-19-related concerns, including access to, perceptions of, and confidence in COVID-19 vaccines and COVID-19-related practices. About 90.7% received the initial vaccine; 77.7% received the booster. About half the participants had received a positive COVID-19 diagnosis by the time of the survey; they were less likely to get the vaccine or booster if they had received the initial vaccine. CONCLUSIONS: Data suggested differences in levels of COVID-19-related concerns and confidence in, or importance of, vaccines when comparing different demographic factors and vaccination practices. The data informs efforts to understand factors affecting vaccine hesitancy among educational professionals.
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An increasing number of cancer subtypes are treated with front-line immunotherapy. However, approaches to overcome primary and acquired resistance remain limited. Preclinical mouse models are often used to investigate resistance mechanisms, novel drug combinations, and delivery methods; yet most of these models lack the genetic diversity and mutational patterns observed in human tumors. Here we describe a series of 13 C57BL/6J melanoma cell lines to address this gap in the field. The Ohio State University-Moffitt Melanoma Exposed to Radiation (OSUMMER) cell lines are derived from mice expressing endogenous, melanocyte-specific, and clinically relevant Nras driver mutations (Q61R, Q61K, or Q61L). Exposure of these animals to a single, non-burning dose of ultraviolet B accelerates the onset of spontaneous melanomas with mutational patterns akin to human disease. Furthermore, in vivo irradiation selects against potent tumor antigens, which could prevent the outgrowth of syngeneic cell transfers. Each OSUMMER cell line possesses distinct in vitro growth properties, trametinib sensitivity, mutational signatures, and predicted antigenicity. Analysis of OSUMMER allografts shows a correlation between strong, predicted antigenicity and poor tumor outgrowth. These data suggest that the OSUMMER lines will be a valuable tool for modeling the heterogeneous responses of human melanomas to targeted and immune-based therapies.
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Línea Celular Tumoral , Melanoma , Animales , Ratones , Línea Celular Tumoral/efectos de la radiación , GTP Fosfohidrolasas/genética , Melanoma/tratamiento farmacológico , Melanoma/genética , Melanoma/patología , Proteínas de la Membrana/genética , Ratones Endogámicos C57BL , Mutación/genética , Proteínas Proto-Oncogénicas B-raf/genéticaRESUMEN
Increasing acknowledgement of climate change (CC) has encouraged various responses, such as education standard mandates. In 2021, New Jersey (NJ) became the first U.S. state to require K-12 CC education across subjects, effective fall 2022. This necessitated introductory science courses on CC to support high school (HS) curricula. Thus, NJ Safe Schools Program (NJSS) created a new course titled, "Introduction to HS Students to CC, Sustainability, and Environmental Justice (EJ)." Given that the COVID-19 pandemic continues (2020-2023 school years) and vaccination coverage varies, this course was developed and approved in an asynchronous online format. Its five modules cover environmental science, CC, natural disasters and extreme weather events, sustainability, including energy conservation and efficiency definitions, and EJ. A 20-question survey included at the end, modified/adapted from a larger nationwide U.S. Student Conservation Association (SCA) survey 2019-2020, examined the perspectives of HS students concerning CC. Selected volunteer NJ HS enlisted students (n = 82/128 finished) to pilot this course February-April 2022. Results such as average scores ≥90% suggested success regarding initial knowledge and awareness gained; for individual modules, two knowledge checks >80% and three knowledge checks >90%. The SCA survey results, overall and by region in NJ, highlighted how most students felt about CC and extreme weather events, plus issues such as EJ. This NJSS introductory course opened in July 2022 for NJ public county secondary school districts and comprehensive HS with approved career-technical education programs, and potentially elsewhere.
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COVID-19 , Cambio Climático , Humanos , New Jersey , Justicia Ambiental , Conocimientos, Actitudes y Práctica en Salud , Pandemias , COVID-19/epidemiología , Instituciones Académicas , EstudiantesRESUMEN
Vaccine hesitancy continues to be prevalent in the United States, especially in relation to the COVID-19 vaccines and its boosters, which have been made increasingly available for public use as the pandemic has progressed. There continues to be concern surrounding the safety and health of secondary or high school education professionals as they transition back to in-person learning and working opportunities. The present study highlights how information dissemination regarding the COVID-19 vaccine has varied among New Jersey secondary or high school teachers throughout the pandemic. The survey was completed online through the PsychData platform by 269 participants between March and July 2022. Participants received the opportunity to complete the survey via email. Afterwards, data were exported and analyzed using Microsoft Excel and SAS 9.4 Analytics Software and stratified by various clinical and demographic-based variables. While trusted agencies and media outlets identified by participants varied, most participants identified the Centers for Disease Control and Prevention (65.4%), primary care providers (37.5%), and state health departments (28.6%) as their top trusted sources for information related to COVID-19 vaccines. Overall, COVID-19 vaccination advocacy and educational efforts should continue across the state of New Jersey and elsewhere, especially as more variants emerge and boosters become available.
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Identifying potential rapid methods to track COVID-19 trends within schools has become a necessity in understanding how to provide both education and maintain health and safety during a pandemic. This study examined COVID-19 trends and sociodemographic information in New Jersey (NJ) schools during the 2020-2021 school year. A database was compiled for this study in Microsoft Excel using various state and federal resources. Data used in the study are a combination of extracted data from weekly NJ Spotlight reports, weekly NJ COVID-19 Activity Level Index (NJ CALI) reports, and reports of school-based outbreaks via the NJ Department of Health (NJDOH). In 2020-2021, in NJ K-12 schools, the NJDOH defined a school-based outbreak incidence as two or more students and/or adult staff with a laboratory-confirmed positive molecular test for COVID-19 based on transmission occurring on campus. Data were organized into six regions across 21 counties within NJ (3-4 counties per region per NJDOH). COVID-19 trends in NJ schools mirrored trends in their districts, i.e., communities, within the state's region; noticeably, there were consistently high trends during the winter holiday season (November 2020-January 2021). The cumulative number of incidences of school-based outbreaks remained relatively low but, nevertheless, increased throughout the 2020-2021 school year. This study recommends increased accessibility to COVID-19 reports for school and public health officials, and in the future for data to be reported to identify rates of transmission of other communicable diseases within K-12 students, and to further reinforce established mandates and other preventative measures in public while traveling during holiday seasons.
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COVID-19 , Adulto , COVID-19/epidemiología , Brotes de Enfermedades , Humanos , New Jersey/epidemiología , Pandemias , Instituciones AcadémicasRESUMEN
Immunotherapy (IT) and targeted therapy (TT) are both effective against melanoma, but their combination is frequently toxic. Here, we investigated whether the sequence of IT (anti-PD-1)â TT (ceritinib-trametinib or dabrafenib-trametinib) was associated with improved antitumor responses in mouse models of BRAF- and NRAS-mutant melanoma. Mice with NRAS-mutant (SW1) or BRAF-mutant (SM1) mouse melanomas were treated with either IT, TT, or the sequence of ITâTT. Tumor volumes were measured, and samples from the NRAS-mutant melanomas were collected for immune-cell analysis, single-cell RNA sequencing (scRNA-seq), and reverse phase protein analysis (RPPA). scRNA-seq demonstrated that the ITâTT sequence modulated the immune environment, leading to increased infiltration of T cells, monocytes, dendritic cells and natural killer cells, and decreased numbers of tumor-associated macrophages, myeloid-derived suppressor cells, and regulatory T cells. Durable responses to the ITâTT sequence were dependent on T-cell activity, with depletion of CD8+, but not CD4+, T cells abrogating the therapeutic response. An analysis of transcriptional heterogeneity in the melanoma compartment showed the sequence of ITâTT enriched for a population of melanoma cells with increased expression of MHC class I and melanoma antigens. RPPA analysis demonstrated that the sustained immune response induced by ITâTT suppressed tumor-intrinsic signaling pathways required for therapeutic escape. These studies establish that upfront IT improves the responses to TT in BRAF- and NRAS-mutant melanoma models.