Translational Research Support Core (TRSC)
Formerly known as the Integrated Health Sciences Facility Core (IHSFC)
The Translational Research Support Core (TRSC) promotes the goals of the Center for Environmental Health and Justice in Northern Manhattan (CEHJNM) by supporting our community partners and existing research projects and by fostering new interdisciplinary research on environmental health and justice (EHJ). The TRSC includes expertise and facilities at Columbia University (CU)’s Lamont-Doherty Earth Observatory (LDEO)—one of the central units of CU’s newly launched Climate School—and CU Irving Medical Center (CUIMC). CUIMC and LDEO provide cost-share support to the TRSC labs, which can therefore offer subsidized member rates
The TRSC offers a comprehensive array of sample collection, field, and laboratory services. The Core is co-directed by Drs. Steven Chillrud and Regina Santella. This joint leadership provides complementary expertise—Dr. Chillrud in environmental matrices and Dr. Santella in biological matrices. The TRSC labs are organized into two sub-cores. Dr. Chillrud leads the sub-core comprising expertise on collection and analysis of environmental samples (e.g., air, dust, water, soil, other matrices) and sensor-based data for external exposure assessment; Dr. Santella leads the sub-core comprising expertise on collection and analysis of biological samples (e.g., animal samples or human blood, urine, saliva, hair, nails, tissues, cells). All six labs perform cutting-edge sample analyses. TRSC services include:
Consultations on sample collection methods and study design;
Processing and archiving environmental or biological samples;
Developing and validating new equipment, including integrated and continuous monitoring for ambient, indoor, and personal monitoring;
Maintain and loan field equipment;
Support community-based research and Community Engagement Core (CEC) activities;
Respond to exposure emergencies;
Train students, staff, junior faculty and fellows;
Provide TRSC lab services;
Coordinate outside services with partner labs.
Biomarkers Lab Leader: Regina Santella, PhD
Trace Metals Lab Leader: Kathrin Schilling, PhD
Biomarkers Lab Manager: Irina Gurvich, MS
Trace Metals Lab Manager: Ronald Glabonjat, PhD
Description of Services
During study design and startup, the IHSFC offers consultations to determine logistics related to biospecimen collection (including personnel training in preparation for the study); preparation of custom protocols for sample collections; and support to biospecimen collection with handling, pre-processing, and aliquoting of study samples. The IHSFC also supports CEHNM’s studies through biorepository management. Its freezer farm provides sample storage and archiving, and prompt sample access and distribution for these studies. It also provides investigators prompt access to existing biobanks that can be used to test new hypotheses; these include a total of 375,000 individual aliquots from 70,000+ participants in 133 human studies of environmental exposures and neurological, cardiovascular, and respiratory diseases, and cancer.
The IHSFC provides analytic support for CEHNM members through two laboratories: a) the Biomarkers Laboratory, which provides an extensive variety of analytical services that range from ELISA assays and microRNA analysis, to library preparations for massive parallel sequencing; and b) the Trace Metals Laboratory, which provides specific consultation to investigators for metals analysis during the planning phases of studies, provides appropriately clean supplies for collecting urine, blood, and other biospecimens, and enables CEHNM members to precisely measure a broad range of metal biomarkers, including As, Pb, Se, Mn, Cd, Fe, and others, in urine, blood, hair, nail and other biological samples. The IHSFC also directs members to services available elsewhere at Columbia University (e.g., at the CTSA or the Comprehensive Cancer Center).
The Biomarkers Laboratory is an extensively utilized shared resource with the Cancer Center, thus fostering integration with other on-campus resources and economy of scale. Services include processing and storage of primarily blood, urine and saliva samples and isolation of DNA for EHS-related and cancer studies. Samples from the 133 studies are stored in the Black building and in a 1,400 ft2 EHS freezer farm located two blocks away that provides expanded sample storage and allows the splitting of individual samples into two locations for enhanced security. The IHSFC freezers (eight -20oC, 24 – 80oC, and four liquid nitrogen freezers) are connected to a Sensaphone alarm system. Sterile blood processing is done in two laminar flow hoods; a CO2 incubator is available for cell culture. Three centrifuges are dedicated to blood processing. Additional equipment includes a Nanodrop, cytospin, 96-well UV, fluorescence, and chemiluminescence readers for DNA concentration determination and ELISA, 96-well plate washer, Qubit fluorometer, Hamilton automated liquid handler and ABI 96-and 384-well real-time thermal cyclers. The bar code system includes two thermal transfer printers and two laser bar code scanners. The lab has recently acquired BioRad CFX96 and CFX384 thermal cyclers that can read fluorescence at two temperatures after each round, which we use for a single-well, multiplexed telomere length assay, which increases the precision of telomere length analysis over traditional two-well assays.
Processing and Storage of Biological Samples
The IHSFC provides initial consultation on all aspects of sample collection and processing, and standard operating procedures are established for each study. Samples are coded to maintain confidentiality using preprinted bar code labels. Blood samples are fractionated according to study-specific protocols, with each vial of each fraction coded for tracking and for their freeze/thaw cycles. The IHSFC also processes and stores urine and oral swabs/saliva samples. As a result of recent member requests, skin microbiome and hair samples are also stored. Costs for these and other services can be found in the website.
Isolation of DNA/RNA
The lab isolates DNA from cells, tissue, and plasma by using various protocols, with routine quality control (QC) determination of the 260/280 ratios. DNA can also be isolated from filter cards and, as an identity check, cross-checked for microsatellite repeats with DNA isolated from aliquoted cells. Stock DNAs are kept in vials but can be prepared in 96-deep-well storage plates, if necessary. Whole-genome amplification is also available. A Qubit is also available for DNA quantitation because several investigators requested this methodology for DNA samples being sent for genotyping. Using commercial kits, the IHSFC now routinely isolates RNA including microRNA from tissue, cells, and plasma for several investigators. Some send samples to the sequencing facilities at the NY Genome Center or Cancer Center; other samples are run in the IHSFC using individual Taqman assays or Taqman Low Density Arrays.
The Biomarkers Lab routinely tests for exposure to polycyclic aromatic hydrocarbons (PAHs) by ELISAs for PAH-DNA and albumin adducts; measures of oxidative stress include ELISAs for urinary 8-oxoG and measurement of protein carbonyls and isoprostanes. Consulting on genotyping platforms is provided, and studies of a limited number of single-nucleotide polymorphisms (SNPs) are done using TaqMan assays in a 384-well format. Larger studies are referred to other campus facilities where Sequenom and Illumina instruments are available.
Biorepository Data Management
Each sample is identified by a unique sample ID that links it with information from questionnaires, examinations, and electronic medical records. After a sample is processed, aliquoted, and stored, a paper record with that sample’s bar code records the date collected and processed, who processed the sample, the sample amount, and how many of each type of aliquot were made using a preset form specific for the particular study. A customized online data entry system, developed by CEHNM’s data management staff, automatically assigns sample location for bulk recording of new sample storage. Database information includes study identifiers, sample ID, aliquot type, volume, location (freezer, shelf, rack, box, and position in box), date received/processed, and technician; tracking of freeze/thaw cycles allows tracking of each vials history. The database stores information on all use and shipment of specimens, and employs a number of internal checks to ensure data integrity. The system uses an interface created in Microsoft Access for dynamic querying and searches, automatic entry of default values, and automation of routine tasks. Data are stored permanently in an MS SQL Server 2014 database, which enforces quality constraints and referential integrity. Access to the data is via an encrypted internet connection, and access is provided for authorized users with valid passwords only. The system is maintained by Mr. Richard Buchsbaum, data manager in the Data FC, and contains information on approximately 375,000 unique biospecimen aliquots. A separate web-based system stores research results, including methods used, person generating the data, and test results. The system allows multiple users to have simultaneous access to the system.
The IHSFC also arranges for investigators to learn how to analyze samples either in their own laboratory or using IHSFC equipment. Laboratory staff provide instruction on the importance of assay specific QC (e.g., blinded, pooled, blanks and duplicate samples) and help investigators set up appropriate batching of samples for analysis (e.g., mixing of exposed and unexposed in each batch).
Trace Metals Laboratory
The Trace Metals Laboratory has been an extraordinary asset since CEHMN inception. It is jointly supported by the CEHNM and the Superfund Research Program. The Trace Metals Lab, adjacent to the Biomarkers Lab, can test blood, urine, hair, nails, serum, and plasma for the presence of As, Pb, Mn, Se, Cd, Cr, Ni, Zn, Cu, Mo, Sb, Pt, U, and W. The lab is equipped with three Perkin-Elmer (PE) Atomic Absorption Spectrophotometers (AAS) and two inductively coupled plasma mass spectrophotometers with dynamic reaction cell (ICP-MS-DRC), which collectively allow for the measurement of a wide range of concentrations of metals, metalloids, and their metabolites in biological samples. The newest of these, a NexION 350s ICP-MS, equipped with SC-4 DX FAST auto-sampler, was recently purchased with an NIH shared instrumentation grant. The AASs include a PE Model AAnalyst800, which has dual interchangeable capability of AAS, flame mode with an S10 auto-sampler, which is less sensitive and allows for the precise measurement of higher concentrations of an array of metals, and graphite furnace mode which provides better sensitivity; and two Graphite Furnace AAS (GFAA), PE Model AAnalyst 600, which can measure different elements at very low concentrations. One ICP-MS is coupled to a PE high-powered liquid chromatography (HPLC) series 200 and is dedicated to As metabolite speciation. Collectively, our instruments can simultaneously determine a large number of elements across a very wide range of concentrations, thus saving considerable time (and sample volumes) needed for analysis. The laboratory also has three PE UV/VIS spectrophotometers, is equipped to prepare dry or wet ash digests of biological samples, and is highly experienced in eliminating sources of contamination.
Sample Collection and Storage
Containers for collecting urine samples are bought as trace metals free or acid washed and bar-coded prior to shipment to investigators. Blood sample tubes for trace metal analysis are also provided by this lab, which routinely obtains large uniform batches that are prechecked for contamination.
Preanalytical Quality Control (QC)
There are many ways in which biological samples can be contaminated—through the use of unsuitable collection instruments, tissue containers, blood collection vials, or contaminated anticoagulants. The pitfalls associated with trace metal analysis are well known to our lab. To prevent the possibility of contamination of blood, urine, or tissue specimens, all specimen collection supplies and plastic-ware for daily work are pre-tested for the metal of interest and acid washed, if necessary.
Standardization and Analytical QC for Metals
The Trace Metals lab participates in QC programs for different elements run by GFAA and ICP-MS. We joined the CDC Blood Lead Laboratory Reference System in 1985. It has been certified by the Occupational Safety & Health Administration. In 1999, the lab joined a unique QC program for urinary As and Pb, run by the Institut de Sante Publique du Quebec; this program provides three "unknowns" five times/ year to laboratories throughout North America/Europe. Another QC program that the lab joined in 2004, run by the same Quebec Institute, is the External Quality Assessment Scheme (QMEQAS). Thrice per year they provide blood, urine, serum, and either hair or nail samples, with 29 different elements of “unknown” concentration, for ICP-MS validation. Since joining, the lab has successfully analyzed most of the samples for elements of interest, including As, Pb, Mn, Se, Cd, Cr, Ni, Zn, Cu, Mo, Sb, Pt, U, and W. To standardize urinary As and other metals, the lab generally used Standard Reference Materials obtained from the National Institute of Standards & Technology (NIST; Dept. of Commerce). Since joining the QC programs run by the Quebec Institute, they decided to use their QC samples because all elements of interest are available in all matrices of interest, and they provide samples on a more regular and timely basis than NIST. However, they still occasionally use NIST standards for some applications.
Analytical Methods for Elements in Blood and Urine
The lab currently utilizes well-established, published GFAA and ICP-MS-DRC methods and sometimes refines them in the laboratory. With regard to past and future work, the following standard methods are used. Blood Pb is analyzed by the GFAA method of Fernandez and Hilligoss, and total urinary As is measured as in Nixon et al. Blood/serum As, Se, Mn and Pb are analyzed by ICP-MS-DRC using modifications of established techniques. The preparation of blood samples according to Stroh was modified based on suggestions from the Quebec Institute, which runs the ICP-MS Comparison Program, while instrument parameters were established using the methods of Pruszkowski et al and Nixon et al. When developing or adapting new GFAA and ICP-MS-DRC methods, the lab relies on the method of additions and the use of appropriate Quebec or NIST standards to validate and standardize the method.
Urinary As metabolites: The lab uses CP-MS-DRC for the speciation of As metabolites in urines. ICPMS-DRC is coupled to HPLC which is used as a detector for As metabolites chromatographically separated on an anion exchange column. Excellent separation by HPLC, coupled with the very low detection limits of ICPMS-DRC, allows us to detect arsenocholine (AsC), arsenobetaine (AsB), monomethyl arsenic (MMA), dimethyl arsenic (DMA), AsIII, and AsV.
Blood As metabolites: The lab successfully adapted the method described above to measure As metabolites in whole blood specimens. Nearly identical to the method for urine, ICP-MS-DRC is coupled to HPLC and used to detect six As metabolites chromatographically separated on anion exchange. Excellent separation power by HPLC, coupled with the very low detection limits of ICP-MS-DRC, allows us to detect AsC, AsB, MMA, DMA, AsIII, and AsV, without online digestion of organic forms and with great precision, even in blood samples with total As concentrations as low as 5 µg/L.