Lan-Huong Tran, PhD Student - Hasselt University

Slides:

Advertisements

Similar presentations

Antibodies Analytical Techniques Utilizing Antibodies: flow cytometry

Advertisements

Ovarian Cancer: How Basic Research Can Lead to New Opportunities for Early Detection and Treatment.

Purification and characterization of EGFP by recombinant Escherichia coli Barney05/10/19.

Surface Plasmon Resonance: antigen-antibody interactions Vamsi K. Mudhivarthi.

4 September, 2006 Chapters Methods: Proteins, Model Systems I.

Engineering receptors and antibodies for biosensors Author: B. Hock, M. Seifert, K. Kramer Presented by: Abu Bakar Md. ISMAIL & Balakrishnan RAMANATHAN.

Detection of Carcinoembryonic antigen with sandwich-biosensor Alexander Pak, Antonina Malozemova, Luiza Niyazmetova, Mukhtar Sadykov, Zeinaf Muradova Nazarbayev.

Biomarkers of ovarian cancer and cysts Reproductive Block 1 Lecture By: Reem Sallam, MD, MSc, PhD.

Construction, Transformation, and Prokaryote Expression of a Fused GFP and Mutant Human IL-13 Gene Sequence Lindsay Venditti, Department of Biological.

Comparison of Serum Human Epididymis Protein 4 with Cancer Antigen 125 as a Tumor Marker in Patients with Malignant and Nonmalignant Diseases J.M. Escudero,

Manufacture of Human Interleukin 13 Protein Using a Prokaryotic Expression System Ryan Rupp, York College of Pennsylvania, Department of Biological Sciences.

Tumor Biomarkers are high lightened Once Again Cancer Facts Cancer etiology Tumor Genetics Tumor MarkersLocation Classification Uses methods of detection.

Antigen-Antibody Interactions : principles and Applications 미생물학교실 권 형 주.

1. 2 What is Click Chemistry? Reactions with the following characteristics: Modular, wide in scope Afford high yields w/o purification Stereospecific.

Tumor markers Present; by Dr. Andalib Isfahan Medical School

CA-125 Cancer Antigen-125 Discovered in l981

吳華席 Hua-Hsi Wu, MD OB/GYN, VGH-TPE Aug 12, 2008

Folding-based Electrochemical Biosensors

It's usually difficult to identify a protein of interest in a Commassie Blue-stained gel of cell extracts Coomassie Blue-stained gel MW stds. Cell extracts.

High-throughput Screening of Soluble Recombinant Proteins Protein Science, 2002, vol 11, YAN-PING SHIH,1 WEN-MEI KUNG,1 JUI-CHUAN CHEN, CHIA-HUI.

Dr. Ranjit Kumar Scientist - I Mahavir Cancer Sansthan Patna, India.

In situ Click chemistry

Introduction and overview of CA19-9 CA19-9 is a tumour associated antigen found in the blood serum. CA 19-9 is also known as the Sialylated Lewis-antigen.

Biomarkers of ovarian cancer and cysts Reproductive Block 1 Lecture Dr. Usman Ghani.

Standad and Novel Biomarkers for Ovarian Cancer 최문선 ∙ 정희윤 1.

Biomarkers of ovarian cancer and cysts Reproductive Block 1 Lecture Dr. Usman Ghani.

Odor-based Sensors for Ovarian Cancer Detection

Figure S1. Production of recombinant NS1 protein

Tumor markers 1111.

GENE EXPRESSION STUDY ON PROTEIN LEVEL

RASSF2DSARAH-GF (b) MST1-F (a) M F G 70 kD a 55 kD b 36 kD

The tNASP protein, a novel diagnostic biomarker for prostate cancer?

The third Annual Conference On Gynecologic Oncology and Prevention Oncology July 20-21, 2017 Chicago, Illinoid USA PPGYNO1 Poster Title.

SURGICAL ONCOLOGY AND TUMOR MARKERS

Lab# 5 Western Blot BCH 462[practical].

Immunoassays are tests that take advantage of the specificity of antibodies to their protein epitotes. Qualitative example: pregnancy test kits, HIV.

5th European Immunology & Innate Immunity Conference

Ahangarzadeh, Sh. *1 Bandehpour, M.1, Kazemi, B.1 , Yarian, F.1

Cancer – a disease of many mutations

SEMINAR 1. Title : Discovery of Protein-Protein Interaction Modulators Using Affinity-Based High-Throughput Screening 2. Speaker : Hyun-Suk Lim (포항공대 (POSTECH))

Progression markers of human epithelial-type ovarian tumors

Large amounts of protein

Affinity and Avidity by: Omar Ammar

A new ligation system for protein conjugation

Globular Protein Made of amino acid chains

Page 1 Site-Specific and Custom Labeling — Creative BioMart.

Target protein Additional file 3. SDS-PAGE showing the degree of purification of D1-26PtxtPL1-27 expressed in E. coli. PtxtPL1-27.

Tumor Marker Tests.

Shahid Bahonar University of Kerman

Enzymes are Proteins with Defined 3D Structures

Engineering yeast to produce proteins for X-ray Crystallography: Heterologous Expression of L. MAJOR proteins in the yeast S. cerevisiae.

Proteon Scientists from Creative Biolabs are willing to provide custom antibody affinity measurement using ProteOn system. The system is a surface plasmon.

Bispecific t cell engager Creative Biolabs possesses unchallenged experience in bispecific T- cell engager (BiTE) synthesis. We elaborately integrate multiple.

Metastasis Models Folder Title: MetModels(NoTP)

Volume 7, Issue 3, Pages (March 2005)

Volume 13, Issue 4, Pages (April 2006)

Practical Virology Lab. (1)

Figure 1: Serum LDH levels elevated significantly in KD and febrile non-KD patients. LDH levels were respectively measured in KD, compared with FC and.

Week 1: Tutorial Outline

Binding of human secretory leukocyte protease inhibitor in uterine cervical mucus to immunoglobulins: pathophysiology in immunologic infertility and local.

Volume 18, Issue 8, Pages (August 2010)

Volume 24, Issue 11, Pages e4 (November 2017)

Ovarian Cancer-Route to Diagnosis

CCL2, Galectin-3, and SMRP Combination Improves the Diagnosis of Mesothelioma in Pleural Effusions Christophe Blanquart, PhD, Fabien Gueugnon, PhD, Jean-Michel.

CLICK CHEMISTRY AND DRUG DISCOVERY

Click Chemistry in Complex Mixtures: Bioorthogonal Bioconjugation

Lab# 5 Western Blot BCH 462[practical].

Drug–Target Association Kinetics in Drug Discovery

Sensitivity and specificity plots of human epididymis protein 4 (HE4) (A and B, respectively) and carbohydrate antigen 125 (CA-125) (C and D, respectively)

Junction Adhesion Molecule Is a Receptor for Reovirus

Presentation transcript:

Selection and characterization of nanobodies for the early detection of ovarian cancer biomarkers Lan-Huong Tran, PhD Student - Hasselt University 4th NanoBio Surfaces and Interfaces in Health Care and Science 2019 Workshop, May 17th 2019, Potsdam, Germany

Ovarian cancer Epithelial ovarian tumors 90-95%, postmenopausal Increases with age Stromal tumors Benign or malignant, age 0-20 Non-aggressive in the adult form Germ cell tumors Benign or malignant, age 10-30 3% of OC Diagnosed in the early stages Tend to grow rapidly

Biomarkers Problems Aims of this project Cancer Antigen 125 (CA125) Human Epididymis protein 4 (HE4) Secretory leukocyte protease inhibitor (SLPI) Progranulin (PGRN) Problems Specificity & sensitivity: still poor at early stage (50% for stage I ) Aims of this project Attempt to develop more efficient biosensor detection strategies based on bioconjugation between Nbs and the biomarkers HE4, SLPI, PGRN To modifify nanobodies (Nbs) targeting biomarker by site-specific functionalization using Expressed protein ligation (EPL)

Expressed protein ligation - EPL

Expressed protein ligation - EPL Copper-Catalyzed Azide-Alkyne Cycloaddition (CuAAC) Click Chemistry

Materials and Methods Human Epididymis protein 4 (HE4) : 13 strains Available strains of Nbs from Vrije Universiteit Brussel (VUB): 35 strains in total Human Epididymis protein 4 (HE4) : 13 strains Secretory leukocyte protease inhibitor (SLPI) : 10 strains Progranulin (PGRN) : 12 strains All strains: Nb-His

An overview of experiment Glycerol stock Pre-culture 37oC Transformation Pre-culture O/N, 37oC Plasmid extraction +IPTG Binding affinity test Expression O/N, 37oC SDS-PAGE on gel 2nd purification, SEC 1st purification, IMAC

Kinetics of Nbs – Binding affinity tests Indirect Elisa Surface plasmon resonance (SPR) KD = koff / kon Epitope mapping koff kon

Results - Elisa A B C

Results – SPR

Coupling of alkynated Nbs to PEG-N3 using click chemistry reaction 15 kDa 25 kDa M: Marker 1: HE4-LEY-CA 2: HE4-LEY 3: SLPI-LEY-CA 4: SLPI-LEY 5: PGRN-LEY-CA 6: PGRN-LEY

Summary Best strains of Nbs from HE4, SLPI and PGRN were chosen The selected nanobodies were successfully site-specifically alkynated at their C-terminus using the Expressed protein ligation (EPL) technique and were coupled to an azidified PEG counterpart using “click”-chemistry.

Thank you for your attention!

Posibilities

Epitope mapping