Nestor Gutierrez Email and Phone Number

Responsible for generating antibodies using B cell culture, bulk single cell sequencing (10X Genomics Chromium X), and hybridoma technology. Continue to optimize new species platforms for VHH nanobodies R&D, generate mRNA for immunization purposes, transfect cell lines (CHO, HEK293), utilize multiplex Next-Generation Sequencing, and discovery new and novel microfluidics machines for increase productivity.

-WT/humanized mice and Rabbit B cell culture: PBMCs purification, single cell technology, gamma irradiation-Bi-specific, vaccine, oncology antibody production -RNA purification, RT-PCR, Gel Imaging, DNA purification, TOPO cloning, sequencing analysis-Various Immunization techniques and bleeds (rats and mice)-ELISA and cELISA and blocking assays-Multi-coloring FACS Sorting [FACSAria II], FACS Calibur, multi-coloring Novocyte, Intelicyte-Automation: Sterile Hamilton and ELISA Hamilton-Hybridoma fusions, subcloning -ClonePix2 usage-ForteBio characterization-ELN notebook data entry, GraphPad, Excel, Word, PowerPoint-Project meeting presentation updates and discussion

• Antibody generation (human, mouse, hamster, and rat)• Prepare immunizations• Hybridoma fusions• Indirect/direct/competitive/blocking/capture ELISA binding assays • HTS FACSCalibur• Limited dilution and ClonePix2 subcloning• ForteBio Octet: Concentration and isotype characterization• Good laboratory/manufacturing practices with tissue culture aseptic techniques• Quartely presentations of data to Hybridoma Discovery Group and monthly project meetings.• General maintenance of TiterTek Plate Reader/Washer, Beckman Coulter Vi Cell, and FACSCalibur.• Generate small scale purification antibody samples using GE HealthCare Protein A/G 96-well columns.• Daily data analysis and concise reports of raw data using Excel, Adobe Reader, Word, E-Notebook, Biobook.• Representative in the Emergency Respone Team. Trained in hazardous, incident, tactics, and fire response.

-Manufacturer and research associate in numerous antigen projects for achieving purified monoclonal antibodies. -Dissected over 300 antigen-boosted mice by performing splenectomies to successfully harvest splenocytes and create hybridoma cells. -Highly applied cell culture techniques to maintain healthy and vibrant cells for biochemical assay testing. -Performed direct ELISA to ensure positive signaling to the recombinant protein. -Utilized western blotting to examine quality assurance of strong human cell line and/or transfected lysate expression. -Ability to work with hazardous chemicals and create buffers. -Trained new employees to master dissection, western blotting, and ELISA techniques. Notable Achievements:-Responsible for manufacturing of over 500 unique purified antigen products: an increase of over 70%.-Simplified the western blot analytical protocol and increased productivity by 45%.-Cost effective idea of reusing laboratory products that saved the company $1000 per month.-Mastered the art of subcloning to acheive a purely monoclonal culture.

-Led out and trained teams of undergraduates in performing microbial physiology experiments, protease activity, and media creation. -Prepared PCR custom template of V. fischeri mutant using General Electric ExoSAP-IT kit.-Aseptically carried out direct cloning of PCR product using TOPO XL PCR cloning kit.-Weekly readings of fluoremeter readings of custom build library bacterial mutants with Molecular Probes EnzCheck® Protease Assay kit.-Autoclaved media and tips. -Weekly meetings with Dr. Patirick Fidopiastis for data analysis and comparison.Notable Achievements:-Journal article published under the Journal of Bacteriology: Manuscript number JB00108-12R1.-Key contributor in discovering relationship of aminopeptidase activity with Vibrio fischeri.

-Performed QC of effluent samples for activated sludge in a tertiary sewage facility using vacuum extractions. -Measured Optical density readings of nitrate and nitrite for effluent samples. -Measured pH, turbidity, and ammonia readings of waste water samples. -Sampled city water distribution stations for detection of Escherichia coli and total coliforms.-Calculated statistical data to meet government standards and regulations. -Aseptically tested bacterial growth of effluent samples in LTB, Brilliant Green, and EC broth media.-Created LTB broth media tubes and nutrient rich agar plates. -Observed daily chlorine readings of creek water beds. -Assisted in city project for measuring carbon dioxide levels in drinking water. -Calibrated pH, turbidity, optical density, and ammonia machines daily. -Recorded temperature levels in incubators and tertiary stations daily. -Assisted in training and supervising newly hired Interns.

-Assisted Clinical/Microbiology Laboratory Scientists in centrifuging and loaded patient samples into automated machines. -Aseptically cultured patient specimens such as stool, urine, spinal/body fluids, wounds, sputum, in aerobic and anaerobic conditions for discovery of pathogens. -Prepared a tremendous amount of Microbiology staining slides such as gram stains, Acid-Fast stains, and Ova and Parasite stains. -Experienced with indirect ELISA kits to diagnosis Influenza A/B, Giardia lamblia, Rotavirus, and Clostridium difficle.-Worked with reference laboratories such as LabCorp and Focus for analysis of patient samples.-Reported and interpreted laboratory results to physicians, nurses, and other health officials. -Helped trained and supervised newly hired employees for quality employee production.