Immunization and serum-collection procedures for producing custom mouse polyclonal antibodies

Our array of Thermo Scientific Pierce Custom Antibody Services includes a customizable option for mouse immunization, serum-collection and titer analysis for mouse polyclonal antibody production.

Highlights

  • Custom mouse antibodies—a reliable protocol for immunizing mice, collecting serum, testing bleeds, reporting titer results and delivering antisera
  • Track and customize project—easily add additional mice to the default protocol; track and manage projects online with OpenProject Tool: you decide which animals to continue or terminate (see Extension below)
  • Full service integration—combine this immunization service with our complete set of services for antigen preparation and antibody purification

5-mouse 80-day protocol

This mouse immunization schedule uses five BALB/c mice for each MAP-peptide, conjugated-peptide antigen, or protein immunogen. Animals are housed individually in separate cages. Injections are subcutaneous (SQ) as emulsions in Complete Freund's Adjuvant (CFA) or Incomplete Freund's Adjuvant (IFA); alternative adjuvants can be used if requested. Continuation of mice is offered on a month-to-month basis. The standard extension protocol includes one boost, one serum collection (0.1mL), and maintenance for one month.

80-day 5-mouse immunization protocol for mouse polyclonal antibody production

Antigen preparation, such as peptide synthesis and/or immunogen conjugation, occurs before Day 0. Injection amounts are given for a conjugated peptide antigen (e.g., KLH), MAP-peptide, or protein immunogen. Protocol days are approximate (± 2 days).

Procedure Protocol Day Description
Control serum collection Day 0 Pre-immune bleed (0.2-0.5 mL per mouse)
Primary injection Day 1 Immunize with 0.10 mg antigen in CFA, SQ 4 sites
1st booster Day 21 Boost with 50 µg antigen in IFA, SQ 4 sites
2nd booster Day 42 Boost with 50 µg antigen in IFA, SQ 4 sites
Test-bleed Day 50 Test-bleed (0.2-0.5 mL per mouse)
ELISA and shipping Day 60 ELISA titration of pre-immune and test-bleed;
(results available online)
3rd booster Day 62 Boost with 50 µg antigen in IFA, SQ 4 sites
Decision Day 64 Verify disposition of mice; decide to continue or terminate
Instruction due date Day 80 Terminal bleed (exsanguination, yielding 0.5-1 mL per mouse)
and termination of animals if instructions not received

Deliverables: 15 vials crude antibody sera

  • 5 vials: 0.1 mL test aliquots pre-immune sera
  • 5 vials: 0.1 mL test bleeds post-inject sera (Day 50)
  • 5 vials: 0.5-1 mL terminal bleeds (exsanguinatory bleed)

Protocol extension

A pre-defined (yet customizable) option is available for extending or continuing the mouse immunization and serum-collection schedule. The purchase decision to continue antibody production can be made at any time before scheduled termination. The terminal exsanguination bleed is included for no cost as part of the default protocol.

One-month extension protocol for mouse antibody production
Procedure Protocol Day Description
1st booster Day 1 Boost with 0.10 mg antigen in IFA, 4 SQ sites
Serum collection Day 14 Bleed (0.1 mL per mouse)

Deliverables (per mouse): 1 vial crude antibody sera

  • 1 vial: 0.1 mL test bleed post-inject sera