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81% of the human proteome on a glass slide.

NEW in v3.1

Expanded content: >20,000 human proteins, encompassing 16,152 genes; more than TWICE the content of other protein arrays

Additional controls: allow for the use of linear regression analysis, radio-labeled sample analysis and the use of different analytical software.

CDI HuProt™ microarray with >20,000 GST fusion proteinsprobed with anti-GST antibody.

Antibody

Testing

PTM

Binding

Binding

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Re-sequenced Content and Expression Th HuProt™ h u m a n p r o t e o m e m i c r o a r r a y v 3.1 c o n t a i n s >20,000 unique, newly sequenced and individually purified human proteins encompassing 16,152 genes (~81% of the human proteome; see fig ure, right) and 124 unique mouse gene symbols. Recombinant proteins are expressed in yeast (S. cerevisiae), purified and printed on glass slides in duplicate, along with control proteins.

HuProt™ v3.1 content can be searched at https://collection.cdi-lab.com/public

Production and Quality Assurance After expression, the N-terminal GST- and His6-tagged pro- teins are purified and printed on glass slides in duplicate, along with control proteins (GST, BSA, Histones, IgG, etc.). Slides are bar-coded for tracking/archiving. Each microarray batch is routinely evaluated by GST staining to insure printing quality and content.

Flexibility is built-in. Th HuProt™ human proteome microarray is available on two types of glass surfaces: PATH™ and SuperE- poxy2™. Focused arrays can be designed out of biomarkers identified within the HuProt™ discovery phase. Efficient (up to 2x7) slide formats can be config ured for a cost-effective and rapid approach to the validation phase.

CDI uses its Arrayjet Ultra Marathon II (photo) non-contact piezoelectric (inkjet) printer to manufacture microarrays with significant performance advantages over contact print methods:

• precise spot morphology (see figure, right)

• large batch sizes for large cohort analysis

• rapid production of custom configured arrays

• reduced inter- and intra-array variability

• greater data reproducibility (see below)

Contact Array Printer NEW Arrayjet Non-Contact

Piezoelectric Printer

Spot Morphology: The piezoelectric “inkjet” process allows the rapid production of high-quality microarray slides time after time, with improved accuracy and reproducibility and excellent spot morphology with even pixel distribution.

P1B

Reproducibility: p=0.95

P3S

Reproducibility: p=0.95 Reproducibility: p=0.89 Reproducibility: p=0.92

High Quality Content and Manufacturing Ensure Reproducibility and Reduces Variability: (A & B) Patient serum sample run in duplicate on two separate HuProt arrays. (C & D) Patient sample from the same blood draw, serum vs plasma run on two separate HuProt arrays. Note: data points are fl escent intensity value of autoantibodies in patient serum bound to individual spotted proteins in the HuProt array.

P2B

P4S

P1

A

P2

A

P3

P

P4

P

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HuProt™ Application Workflow Examples

Body Fluid Biomarker Profiling

(serum, plasma etc.)

HuProt Proteome Microarray Less than 5 µl body fluid/test

Antibody is incubated with HuProt™

and binds with one (or more) proteins

Patient body fl id is incubated with HuProt™

and autoantibodies bind to proteins

expected

target

off-target

cross-reactivity

Microarray washed and incubated with

a fl orescent secondary antibody

Microarray washed and incubated with

isotype-specifi fl orescent secondary

antibodies (anti-IgG, IgM, IgA)

Antibody-protein binding is analyzed

HuProt™ allows for a quantum leap in assessing the

quality of antibodies used in research and manufacturing

Large experimental / control population studies

can reveal novel biomarkers (see overleaf)

White Paper: cdi-lab.com/High-Spec_White_Paper.shtml Video Demo: cdi-lab.com/Video.shtml

expected target

statistically signifi ant

cross reactivity

(if there is none, the Ab is monospecifi )

Specificity

and specifi ty is assessed

rela

tive s

ign

al in

ten

sity

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A Closer Look: 2-Phase Biomarker Discovery Workflow - powered by HuProt™

Summary

• Cost-effective, rapid tool for early, companion and predictive diagnostics

• Can be performed as a CDI service of by clients in-house

• Step-by-step training and continued support

• Rapid turnaround and comprehensive reporting

Phase 1 - Discovery

After bioinformatic analysis, profiles from both groups are compared and candidate biomarkers are identified.

Samples of patients with the disease of interest as well as normal samples are tested on HuProt™. Typically, 50-100 samples for each category are used.

Phase 2 - Validation

The biomarker candidates from the discovery phase are used to generate focused arrays. 14-64 arrays can be printed per slide dramatically reducing cost and allowing much larger cohorts of patient samples to be tested.

Video Demo: cdi-lab.com/Video.shtml

The robust, rapid validations of biomarkers eliminates problems associated with data overfitting.

The validated biomarkers are immediately transfer- able to ELISA-based assays or other commercial assay platforms

Custom Discovery Services Using the HuProt™ Microarray, CDI can do the work for you so you can concentrate on your research. We offer expertise in high-throughput design and provide detailed data analysis.

Developed an antibody? The specificity will be critical to the success of your research. High-Spec® Antibody Cross-Reactivity Testing is offered to assess cross-reactivity to the largest num- ber of human proteins - HuProt™ - BEFORE you publish. Read the new WHITE PAPER on commercial antibody specificity.

Access it here: cdi-lab.com/High-Spec_White_Paper.html

Call us toll-free at 844-539-6296 or email us at info@cdi-lab.com for more information or to obtain a quote and begin a project.

HuProt™ Human Proteome Microarray Literature Citations

Cox E et al. (2017) Global Analysis of SUMO-Bind- ing Proteins Identifies SUMOylation as a Key Reg- ulator of the INO80 Chromatin Remodeling Com- plex. Mol Cell Proteomics 16, Mar 2. doi: 10.1074/ mcp.M116.063719.

Zhaoshou Y et al. (2017) A human proteome array approach to identifying key host proteins target- ed by Toxoplasma kinase ROP18. Mol Cell Pro-

teomics 16, doi:10.1074/mcp.M116.063602mcp. M116.063602

Barry G et al. (2017) The long non-coding RNA NEAT1 is responsive to neuronal activity and is associated with hyperexcitability states. Na-

ture Scientific Reports 7, Article number: 40127 doi:10.1038/srep40127

Shi L et al. (2016) Application of high-throughput protein array in clinical screening for tumor mark- ers. Int J Clin Exp Med 9:8529-8535.

Iyama S et al. (2016) Drebrin: A new oncofetal biomarker associated with prognosis of lung ade- nocarcinoma. Lung Cancer 102:74-81.

Cheng X et al. (2016) Proteomic identification of the oncoprotein STAT3 as a target of a novel Skp1 inhibitor. Oncotarget DOI: 10.18632/oncotar- get.13153.

Hu CJ et al. (2016) Identification of Novel Biomark- ers for Behcet Disease Diagnosis Using HuProt Ar- ray Approach. Mol Cell Proteomics 16:147-156.

Wang Y et al. (2016) A nuclease that mediates cell death induced by DNA damage and poly(ADP-ri- bose) polymerase-1. Science 354 Oct 7 DOI: 10.1126/science.aad6872.

Li H et al. (2016) Penetrance of Congenital Heart Disease in a Mouse Model of Down Syndrome Depends on a Trisomic Potentiator of a Disomic Modifier. Genetics 203:763-70.

Ogishi M et al. (2016) Delineation of autoantibody repertoire through differential proteogenomics in hepatitis C virus-induced cryoglobulinemia. Sci

Rep 6:29532.

Yang L et al. (2016) Identification of serum bio- markers for gastric cancer diagnosis using a hu- man proteome microarray. Mol Cell Proteomics

15:614-23

Syed P et al.(2016) Autoantibody Profiling of Gli- oma Serum Samples to Identify Biomarkers Using Human Proteome Arrays. Sci Rep 5:13895. doi: 10.1038/srep13895.

Ainscough JS et al. (2015) Interleukin-1β Process- ing Is Dependent on a Calcium-mediated Interac- tion with Calmodulin. J Biol Chem 290:31151-61.

Zhang K et al. (2015) The C9orf72 repeat expan- sion disrupts nucleocytoplasmic transport. Nature

525:56-61.

Zhang HN et al. (2015) Systematic identification of arsenic-binding proteins reveals that hexokinase-2 is inhibited by arsenic. Proc Natl Acad Sci USA

112:15084-9.

Hu J et al. (2015) Systematic Prediction of Scaffold Proteins Reveals New Design Principles in Scaf- fold-Mediated Signal Transduction. PLoS Comput

Biol. 2015 11: e1004508.

Cox E et al. (2015) Identification of SUMO E3 ligase-specific substrates using the HuProt hu- man proteome microarray. Methods Mol Biol

1295:455-63.

Hu C et al. (2015) Autoantibody Profiling on Hu- man Proteome Microarray for Biomarker Discov- ery in Cerebrospinal Fluid and Sera of Neuropsy- chiatric Lupus. PLoS One 10:e0126643.

Liu S et al. (2014) Characterization of monoclonal antibody’s binding kinetics using oblique-inci- dence reflectivity difference approach. MAbs

7:110-119.

Jung JG et al. (2014) Notch3 interactome analy- sis identified WWP2 as a negative regulator of Notch3 signaling in ovarian cancer. PLos Genet

10:e1004751.

Fan Q et al. (2014) Identification of proteins that interact with alpha A-crystallin using a human pro- teome microarray. Mol Vis 20:117–124.

Deng RP et al. (2014) Global Identification of O-GlcNAc Transferase (OGT) Interactors by a Human Proteome Microarray and the Construc- tion of an OGT Interactome. Proteomics 14: 1020-30. doi: 10.1002/pmic.201300144. Epub 2014 Mar 25.0.

Ma TM et al. (2014) Serine Racemase Regulated by Binding to Stargazin and PSD-95: Potential NMDA-AMPA Glutamate Neurotransmission Cross-talk. J Biol Chem 289:29631-41..

Barry G et al. (2013) The long non-coding RNA Gomafu is acutely regulated in response to neu- ronal activation and involved in schizophrenia-as- sociated alternative splicing. Molec Psychiatry

19:486-94

Lee YI et al. (2013) Protein microarray character- ization of the S-nitrosoproteome. Mol Cell Pro-

teomics 13:63-72.

Chen Y et al. (2013) Bcl2-associated Athanogene 3 Interactome Analysis Reveals a New Role in Mod- ulating Proteasome Activity. Mol Cell Proteomics

12:2804-19.

Donnelly CJ et al. (2013) RNA toxicity from the ALS/FTD C9ORF72 expansion is mitigated by an- tisense intervention. Neuron 80:415-28.

Fan B et al. (2013) A human proteome microarray identifies that the heterogeneous nuclear ribonuc- leoprotein K (hnRNP K) recognizes the 5’ terminal sequence of the hepatitis C virus RNA. Mol Cell

Proteomics 13:84-92.

Tarrant MK et al. (2012) Regulation of CK2 by phosphorylation and O-GlcNAcylation revealed by semisynthesis. Nat Chem Biol 8:262-9.

Jeong JS et al. (2012) Rapid identification of monospecific monoclonal antibodies using a hu- man proteome microarray. Mol Cell Proteomics

11:O111.016253.

Huang Y et al. (2012). Global tumor protein p53/ p63 interactome: making a case for cisplatin che- moresistance. Cell Cycle 11:2367-79.

Hu CJ et al. (2012). Identification of new auto- antigens for primary biliary cirrhosis using hu- man proteome microarrays. Mol Cell Proteomics

11:669-80.

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