UNC Biochemistry Core Facilities and Resources

 

 The biochemistry cores offer resources to investigators working on the molecular analysis of genes, proteins, peptides, cells and tissues.  Analytical approaches include mass spectrometry, NMR spectroscopy, x-ray crystallography and bioinformatics.

Immunology


CFAR Virology, Immunology & Microbiology Core 
provides support for research among CFAR (Center for AIDS Research) investigators and training in sample preparation and/or the performance of immunologic procedures.

Research applications: Sample processing and storage, HLA-typing, CD4 counts, measurement of soluble biomarkers (ELISA, Luminex platform, ELISPOT), flow cytometry for phenotyping and functional analysis of immune cell populations; antibody analysis.

Instrumentation available: Luminex, MagPix Instruments, 6-8 color flow cytometry instruments –FACS Canto (BD), FACS Calibur (BD), ELISA reader, multiple PCR instruments.

Eligible Users: UNC infectious disease clinicians, CFAR–affiliated researchers, biotech companies, other clinicians and researchers interested in infectious diseases.

 

Antibody Core Facility can help investigators produce monoclonal antibody-producing hybridoma cell lines. The core has partnered with an outside vendor (Precision Antibody) to work with investigator labs to develop hybridoma lines. Once developed, the core can take possession of these lines, store them for the investigators, and carry out large scale production and purification of the antibodies (multi-milligram). The facility also can produce large amounts of crude antibody in vitro for investigators (conditioned media) and purifies both polyclonal and monoclonal antibodies from supplied tissue culture media or ascites. If the Antibody Core is able to extract the antibody sequence from a hybridoma cell line, the Protein Expression and Purification Core – located in the same building – can produce it recombinantly using mammalian expression systems. This facility can also help you to produce protein antigen(s) you may require for your hybridoma project.

Operation: We help the investigator lab think through and structure their project, coordinate its execution and produce the resulting custom antibodies in our facility once the hybridoma lines are established. We help them produce antigens as needed via the Protein Expression and Purification Core, connect them with our commercial partner precision antibody, and try to facilitate the process in any way we can

Research applications: Generation of hybridoma cell lines from mice, rats and hamsters to produce monoclonal antibodies that specifically recognize an antigen(s) of interest. Design and production of protein antigens in the Protein Expression and Purification Core, which is located in the same facility. Production of antibody from of hybridoma cell culture. Purification of polyclonal and monoclonal antibodies from tissue culture media or ascites. Conjugation of purified antibodies to a variety of things like fluorescent tags, small molecules like biotin, other proteins, and chromatography media. Tissue culture services are also offered- examples: adapting cell lines to various medias and suspension culture.

Instrumentation available: Animal work is done with our commercial partner (Precision Antibody). Notable equipment – a Biotek plate washing robot which enables high throughput ELISA screening; several western blot devices that enable screening of up to 50 sera per blot. Copious amounts of 37C CO2 regulated incubator space (including shaking platforms). We also are co-localized with the protein expression and purification core, providing access to all their resources.

Training resources: Individualized training (fee-for-service) of users for some aspects of our service is offered, please contact for more detail.

Eligible users: Researchers at UNC, other academic institutions as well as commercial laboratories.

 

Mass Spectrometry


The Biomarker Mass Spectrometry Facility provides expertise for qualitative and quantitative analysis of small molecular weight biomarkers including nucleic acids, peptides, and metabolites. In addition to LC/MS/MS analyses, we offer HPLC with UV detection and trace elemental analysis by ICP-MS. Areas of focus for the facility have included DNA and protein adducts as markers of chemical exposure, oxidative stress, and endogenous DNA damage, characterization of chemical reaction products, identification and quantitation of metabolites, multi-element analysis and arsenic speciation. The services and analyses performed within the facility enable UNC investigators to study the molecular mechanisms of environmentally based diseases as well as the relationships between genetic and environmental factors.

Research applications: Targeted quantitation of biomarkers of exposure and effect including DNA adducts, protein adducts, DNA-DNA and DNA-protein crosslinks, metabolites, and metals; Unbiased differential mass profiling; HPLC purification of reaction mixtures; Molecular weight and structural characterization of synthetic standards.

Instrumentation: Sciex 6500 triple quad mass spec with Eksigent nanoLC; Thermo TSQ Quantum Ultra triple quad mass spec with Waters NanoAcquity UPLC; Thermo TSQ Quantum Ultra triple quad mass spec with Waters Acquity UPLC; Agilent 6520 Accurate-Mass Quadrupole Time-of-Flight mass spec with Agilent 1200 Rapid Resolution LC; Agilent 7500cx ICP-MS Inductively Coupled Plasma mass spec with Agilent 1260 Infinity Bioinert HPLC; Agilent 1200 HPLC for offline purification and fraction collection.

Resources: Full service sample analyses as well as training for independent operation; Consultation for proposal submission and/or project feasibility; Method development; Data interpretation and generation of experimental sections and figures for publications. Fees are based on hourly usage of instrumentation and staff time, so costs correlate with the degree of effort required and the duration of analyses.

Eligible users: UNC researchers as well as other academic investigators and commercial clients when feasible.

 

The Department of Chemistry Mass Spectrometry Core Laboratory specializes in small molecule mass spectrometry analysis. Our Core aims to provide the highest quality mass spectrometric services possible to both campus and surrounding Research Triangle area members. We offer both independent operator and full-service mass spectrometry workflows.  Our primary research focuses are organic and inorganic synthetic molecules, relative and absolute quantitation mass spectrometry, and trace metal analysis.

Research Applications: quantitative mass spectrometry, liquid chromatography separations, structural elucidation, MS/MS & MSn fragmentation mass spectrometry, complex mixture analysis, molecular formula confirmation, high resolution and accurate mass analysis, & trace metal determination/quantitation.

Instrumentation: ThermoFisher Exactive GC, ThermoFisher LTQ FT with a Waters Acquity H-Class UPLC system, PESciex 3000 with a Shimadzu LC and CTC autosampler, ThermoFisher Element XR equipped with ESI Autosampler and Teledyne Cetac Excite 193 Laser Ablation system, Agilent 1100 liquid chromatograph system, CEM Mars Microwave System

Resources: Individualized user training for undergraduate, graduate, and post-doctoral scholars of the Department of Chemistry and UNC Campus community. Walk-up access is available to all trained users. Full-service sample analysis. Consultation for project development. Method development and advising. Data interpretation. Grant application assistance.

Eligible Users: All researchers at UNC-CH, affiliated universities, non-affiliated universities, private industry/commercial clients


UNC Michael Hooker Proteomics Center provides researchers with state-of-the-art analysis of proteins from tissues, cells and other biological samples. The Center is also a resource for hands-on-training, education, and consultation in modern proteomics techniques. The Center offers an array of sample preparation and instrumentation services, as well as method development and in-depth consultation to help researchers optimize their experimental design.

Research applications: Identify purified proteins and co-immunoprecipitated proteins. Identify proteins in from complex mixtures and quantify differences in protein abundance using labeled (SILAC, TMT) or label-free approaches. Identify of post-translational modifications such as acetylation, ubiquitylation, phosphorylation and sulfhydryl modifications. Determine the exact mass of purified peptides and proteins.

Instrumentation available: Thermo Easy nLC 1000; Thermo QExactive HF mass spectrometer; Waters nanoAcquity; Thermo LTQ-Orbitrap Velos mass spectrometer; Sciex 5800 MALDI-TOF/TOF mass spectrometer

Training resources: Individualized training is available for the use of the MALDI-TOF/TOF instrument and data analysis using Protein PilotTM software.

Eligible users: Researchers at UNC and other academic institutions or commercial laboratories.

 

Metabolomics


The Metabolomics Laboratory is a collaborative resource for global and targeted metabolite profiling from a wide range of biological samples.

Research applications: NMR analysis and absolute quantitation on nearly 300 metabolites including, amino acids, organic acids, amines, nucleic acids, carbohydrates, drug related metabolites, etc. from a variety of biological samples (such as urine, serum, CSF and saliva) cell and tissue extracts. Based on research needs, perfused bioreactor studies are available for intact tissue samples or cell suspensions. Concentrations ranging from micromolar up through millimolar levels can be detected. A microcoil NMR probe requiring only 15 µl injections that can be recaptured after analysis, permits concentration of samples thereby pushing detection limits to low micromolar levels, with samples amounts of just 5 µl - typical of mouse plasma metabolomic studies. Typical output for users is a principal component analysis of PC1 versus PC2 with a list of biomarkers derived from the loadings plot.  For <15 media, plasma, or urine samples plan for a week turn around.

Instrumentation available: Varian INOVA 600MHz NMR 
2 channels with pulsed field gradients, and waveform generators, used with Varian 5mm PFG ID probe
or Protasis 10µl HC capillary NMR probe. A Varian SMS 100 robotic sample changer for can queue up 100 samples in 5mm NMR tubes for automated analysis. A Protasis High Throughput fluidics system for the automated injection of small samples (down to 15 µl volume) has three temperature controlled drawers and can accommodate up to 2 x 96 well-plates or 200 of vials per drawer (600 samples total) for automated fluidic injection of samples into the Protasis 10µl HX capillary NMR probe.

Eligible users: Researchers at UNC and other academic institutions or industrial laboratories.

Nuclear Magnetic Resonance (NMR)


The UNC Biomolecular NMR Laboratory supports research into the structure and dynamics of biological molecules.
Research applications: Macromolecular structure and dynamics. Cryoprobes are available for small molecules and biological applications, including small molecule and natural product NMR.

Instrumentation available: Bruker Avance 500 MHz with cryoprobe, Bruker Avance 600 MHz with cryoprobe, Varian Inova 700 MHz with coldprobe.

Planned purchases: Convert Inova 700 to Bruker Avance 700 MHz with cryoprobe, Bruker Avance 850 with cryoprobe and 24 slot sample changer.

Training resources: one-on-one training of new users, consultation with prospective users to determine whether NMR will be useful in their research, and consultation in designing experiments. NMR course BIOC663 open to graduate students.

Eligible users: Any researchers at UNC or other academic institutions, and in industry.

 


School of Pharmacy NMR
provides a resource for small molecule and natural product NMR related to drug discovery.

Research applications: Small molecule and natural product NMR. Routine 1D and 2D 1H and 13C spectra, as well as NMR on 31P, 15N, and other nuclei. Macromolecular NMR experiments are available on the 500 MHz instrument.

Instrumentation available: Varian Inova 500 and Inova 400 spectrometers. A second 400 MHz (Varian 400MR) instrument is located at the satellite laboratory in the Genetic Medicine Building. Multiple probes are available on the 500, including a 3 mm probe for low mass samples.

Training resources: Frequent training sessions (from 1-4 trainees) scheduled based on demand. Free consultations for prospective users.

Eligible users: Any researchers at UNC or other academic institutions, or industry.

 

The Metabolomics Laboratory (by NMR) is a collaborative resource for global and targeted metabolite profiling from a wide range of biological samples.

Research applications: NMR analysis and absolute quantitation on nearly 300 metabolites including, amino acids, organic acids, amines, nucleic acids, carbohydrates, drug related metabolites, etc. from a variety of biological samples (such as urine, serum, CSF and saliva) cell and tissue extracts. Based on research needs, perfused bioreactor studies are available for intact tissue samples or cell suspensions. Concentrations ranging from micromolar up through millimolar levels can be detected. A microcoil NMR probe requiring only 15 µl injections that can be recaptured after analysis, permits concentration of samples thereby pushing detection limits to low micromolar levels, with samples amounts of just 5 µl - typical of mouse plasma metabolomic studies. Typical output for users is a principal component analysis of PC1 versus PC2 with a list of biomarkers derived from the loadings plot.  For <15 media, plasma, or urine samples plan for a week turn around.

Instrumentation available: Varian INOVA 600MHz NMR 
2 channels with pulsed field gradients, and waveform generators, used with Varian 5mm PFG ID probe
or Protasis 10µl HC capillary NMR probe. A Varian SMS 100 robotic sample changer for can queue up 100 samples in 5mm NMR tubes for automated analysis. A Protasis High Throughput fluidics system for the automated injection of small samples (down to 15 µl volume) has three temperature controlled drawers and can accommodate up to 2 x 96 well-plates or 200 of vials per drawer (600 samples total) for automated fluidic injection of samples into the Protasis 10µl HX capillary NMR probe.

Eligible users: Researchers at UNC and other academic institutions or industrial laboratories.

Peptide Synthesis


High Throughput Peptide Synthesis and Array Facility
provides researchers with high quality consultation and peptide synthesis services including production, purification, and characterization of polypeptides and custom designed peptide arrays.

Research applications: Synthetic peptides containing standard, post-translationally modified (e.g., acetylation, methylation, phosphorylation) and/or unnatural/nonstandard amino acids and fluorescent tags (e.g., 6-Cl-Trp, FAM-, TAMRA-) can be synthesized for a wide range of biological and biophysical applications, including FP, ITC, NMR, X-ray crystallography, mass spectrometry (with or without stable isotopes such as 2H, 13C, 15N), enzyme inhibition/competition assays, protein-protein interaction studies, cell permeable peptide reagents, antigen/antibody production including synthesis of Multiple Antigenic Peptides (MAPs), and high density peptide arrays.

Instrumentation available: Symphony multiple peptide synthesizer, PS-3 peptide synthesizer, Waters preparative, semi-preparative and analytical HPLC instruments. Microbalance equipped with static charges neutralizing device.

Training resources: Consultation with users to determine whether synthetic peptides and peptide chemistry may be useful in their research, and consultation in designing their experiments.

Eligible users: Any researcher at UNC or other academic institutions, and in industry.

Structural Biology


The Macromolecular Interactions Facility (Mac-In-Fac) provides instrumentation and resources for biophysical characterization of biological macromolecules and their interactions with cognate ligands.

Research applications: Determination of solution molecular weight, oligomeric state and association/dissociation constants, sedimentation, diffusion and frictional coefficients, hydrodynamic and RMS radius, second virial coefficient, on  and off rates (kon and koff), stoichiometry, equilibrium binding constants and other thermodynamic parameters (∆G, ∆H and ∆S) of bimolecular interactions, secondary structure, melting profile and melting temperature (Tm).

Instrumentation available: Surface plasmon resonance-based biosensors; analytical ultracentrifuges with absorbance, interference, and fluorescence detection systems; a spectrofluorometer; differential scanning and isothermal titration calorimeters; circular dichroism spectropolarimeters, static and dynamic light scattering instruments, and a fluorescence microplate reader.

Training resources: One-on-one training of new users, consultation with prospective users to determine whether techniques available at the Mac-In-Fac will be useful in their research, and consultation in experimental design, data analysis and interpretation. The Macromolecular Interaction course BIOC662 (cross listed as CHEM 735), taught by Ash Tripathy each spring, is open to graduate students. The course includes both theory and  practicals.

Eligible users: Any researchers at UNC or other academic institutions, and in industry.

 


Macromolecular X-Ray Crystallography Core Facility
(MX) provides support and infrastructure necessary to enable individual investigators of any experience level to initiate and complete a macromolecular structural biology project.

Operation: Three general ways users interact with us that can be adjusted at any time: (1) Facility can be an extension of your lab, with trained users operating independently. (2) We collaborate with your group, mentoring your personnel, helping with experimental design and analysis, and aiding with grant writing and manuscript preparation (3) We independently tackle your project.

Research applications: High-Throughput protein crystallization screening, monitoring, and crystal diffraction testing as well as data collection, crystal structure determination and analysis. Sample preparation may be done in the Protein Expression and Purification Core, which is located in the same facility.

Instrumentation available: Crystallization - Wyatt plate based (384) DLS instrument for validation of sample quality; Rigaku Phoenix liquid handling robot for 96 well crystallization plate setup; an Alchemist II liquid handler for custom 96 and 24 well crystallization screen generation; a Minstrel HT imager for automated monitoring of crystallization trials;  Jansi UVEX UV florescence microscope for inspection of crystallization experiments as well as multiple Leica stereo light microscopes with cameras for manual plate inspection and crystal harvesting/manipulation. Diffraction - Rigaku Micromax 007HF generator: Port one equipped with an R-Axis IV++ detector optimized for crystals with a long cell axis; Port two is equipped for automated diffraction screening with an ACTOR robotic system and Saturn 944+ CCD detector. Both beamlines are equipped with an X-stream 2000, enabling cryo-crystallography. A Rigaku LN40 liquid nitrogen generator is available for 24/7 on-demand LN2.

Training resources: Individualized training (fee-for-service) of users in any aspect of a crystallography project.

Eligible users: Researchers at UNC, other academic institutions as well as industrial laboratories; UNC LCCC members receive discounted fees for certain services.

 

The Protein Expression and Purification Core specializes in the production of pure, functional proteins for structural, biophysical, and biochemical studies. It is specifically designed as a “front-end” interface to other components of the Center for Structural Biology. In addition, the services of this core are also applicable to other research areas, including antibody and antigen production and the manufacture of protein chips.

Research applications: Production of purified protein suitable for biochemical, biophysical, and structural studies, the latter of which may be completed in the Macromolecular x-ray Crystallography Core located in the same facility. Produce purified protein suitable for antibody production, which may be done in the Immunology Core Facility located in the same facility. Generation of stably transfected, expression optimized mammalian cell lines. Their uses include: (1) production of your protein of interest in high amounts over multi-week timescales. (2) The cells themselves are a reagent for certain cell based assays.

Instrumentation available: The Core possesses the resources needed for growth and maintenance of large volumes of E. coli (temp. controlled shakers and a 10L fermenter), insect cells (baculovirus mediated, Sf9 and Hi-five cell lines), and mammalian cells (including HEK 293s and CHOs, both suspension and adherent.). A variety of FPLC systems for automated, multi step, multi sample purifications as well as more traditional single step, single sample purifications and method devlopment. We also have equipment that enables gel electrophoresis/western blotting and imaging, small-volume (.5ml to 400ml) as well as large volume (<10L) sample concentration, spectrophotometric analysis (NanoDrop), PCR, cloning, electroporation of E. coli, as well as limited proteolysis analysis of your protein sample.

Training resources: Individualized training (fee-for-service) in methods of protein expression and purification as well as instrumentation operation.

Eligible users: Researchers at UNC, other academic institutions as well as industrial laboratories; UNC LCCC members receive discounted fees for certain services.

The R. L. Juliano Structural Bioinformatics Core Facility provides researchers with state-of-the-art consultation and collaboration in research studies requiring computational structural biology methods.

Research applications: Bioinformatics analysis includes multiple sequence alignments, phylogenetic analysis, and ancestral sequence predictions. Protein structure analysis includes protein fold determination, structure 
superimposition and comparison, homology modeling, active site identification, mutation effects analysis, electrostatic surfaces, solvent accessibility calculations, subunit contact analysis, molecular dynamics calculations, free energies of binding, and docking of small molecules.

Instrumentation available: Researchers utilize hardware and software resources provided by ITS Research Computing. Available software packages include but are not limited to PyMOL, BLAST, T-Coffee, Discovery Studio, VMD, Gromacs and Amber.

Training resources: Training workshops on the following topics: (1) Protein Data Bank (PDB), (2) PyMOL, and (3) Protein fold recognition and homology modeling. Individualized consultation and collaboration on research studies requiring computational structural biology methods.

Eligible users: Researchers at UNC and other academic institutions or industrial laboratories.

Protein Expression

The Protein Expression and Purification Core specializes in the production of pure, functional proteins for structural, biophysical, and biochemical studies. It is specifically designed as a “front-end” interface to other components of the Center for Structural Biology. In addition, the services of this core are also applicable to other research areas, including antibody and antigen production and the manufacture of protein chips.

Research applications: Production of purified protein suitable for biochemical, biophysical, and structural studies, the latter of which may be completed in the Macromolecular x-ray Crystallography Core located in the same facility. Produce purified protein suitable for antibody production, which may be done in the Immunology Core Facility located in the same facility. Generation of stably transfected, expression optimized mammalian cell lines. Their uses include: (1) production of your protein of interest in high amounts over multi-week timescales. (2) The cells themselves are a reagent for certain cell based assays.

Instrumentation available: The Core possesses the resources needed for growth and maintenance of large volumes of E. coli (temp. controlled shakers and a 10L fermenter), insect cells (baculovirus mediated, Sf9 and Hi-five cell lines), and mammalian cells (including HEK 293s and CHOs, both suspension and adherent.). A variety of FPLC systems for automated, multi step, multi sample purifications as well as more traditional single step, single sample purifications and method devlopment. We also have equipment that enables gel electrophoresis/western blotting and imaging, small-volume (.5ml to 400ml) as well as large volume (<10L) sample concentration, spectrophotometric analysis (NanoDrop), PCR, cloning, electroporation of E. coli, as well as limited proteolysis analysis of your protein sample.

Training resources: Individualized training (fee-for-service) in methods of protein expression and purification as well as instrumentation operation.

Eligible users: Researchers at UNC, other academic institutions as well as industrial laboratories; UNC LCCC members receive discounted fees for certain services.

 

Analytical and Physical Chemistry

The Nanomedicines Characterization Core Facility (NCore) provides  comprehensive characterization of novel nanomedicines and nanomaterials.  Services include physicochemical assays (ICP-MS, Nanosight), comprehensive physicochemical characterization studies, and bioassays characterization services.

Research applications: Physicochemical characterization (PC) of nanomaterials offers a comprehensive body of information that can be used to predict behavior of the nanomaterials in biological environments both in vitro and in vivo.  This information is essential in order to exercise tight control over inter-batch variations and maintain reproducibility of nanomaterials critical for their applications.

N-Core offers PC of various classes of nanomaterials such as:

  • Polymer conjugates
  • Polymeric micelles
  • Liposomes
  • Nanogels
  • Polyion complexes of small drugs and biomacromolecules (proteins, DNA, and RNA)
  • Inorganic/metal nanoparticles
  • Bio-derived nanoparticles such as exosomes with protein and nucleic acid cargo

 Eligible users: Any researcher at UNC or other academic institutions, and industry.