Phospho Antibody Custom Service

Antigen Preparation

Non-modified peptide synthesis (10mg, within 15AA, >90% purity) and conjugation

1-2 weeks

Phosphorylated peptide synthesis (15mg, within 15AA, >90% purity) and conjugation

1-2 weeks

Animal Immunization (2 New Zealand white rabbits)

Modified peptide-KLH standard immunization: 4 doses with optional 1-2 booster immunizations

6-8 weeks

Blood Sampling

2 rabbits exsanguinated → ~30-50 ml serum/rabbit

2 weeks

Antibody Purification

Affinity purification using phospho-specific selection against both phosphorylated and non-phosphorylated peptide variants

1-2 weeks

Purified Antibody Assays

ELISA titer determination and delivery of all purified antibodies (minimum 2 mg)

1-2 weeks

Total

3-4 months

Antigen Preparation

Modified peptide synthesis and conjugation (10mg KLH + 5mg BSA + 5mg peptide)

4

Unmodified peptide synthesis and conjugation (5mg BSA conjugate + 5mg peptide)

FITC conjugation of modified peptide (2mg FITC-labeled)

Animal Immunization (2 rabbits)

Primary immunization: 4 doses

Optional boosters: 1-2 doses

ELISA titer monitoring

Delivery of hyperimmune sera from 2 rabbits for client verification

2

Isolation of splenocytes followed by antigen-reactive B cell selection (10-plate scale)

Selection of the highest-titer rabbit for B cell isolation

2

Two-phase high-throughput antibody functionality screening

Primary screen: Phosphopeptide plates (enrichment of phospho-specific B cells)

Counter-screen: Non-phosphopeptide plates (exclusion of non-specific binders)

Output: Delivery of all antigen-positive hybridoma supernatants for client verification

2

Large-scale antibody manufacturing and purification (3 runs)

Three top-performing sequences were selected for antibody production, with 100 μg of each antibody prepared. The three antibody sequences and remaining peptides will be provided.

2

Total

3-4 months