MACS Matchmaker

Fc-receptor capture — protocol

For Fc-effector profiling and antibody PK / half-life screening, lino Biotech supplies pre-conjugated Fc-receptor–oligo adapters that hybridise directly to any Oligo|PEG chip. No off-chip conjugation, no IMAC step — flow the adapter, then flow the antibody analyte. Two ready-to-use adapters cover the common questions: a multiplexable FcγR adapter (FcγRI, FcγRIIa, FcγRIIb, FcγRIIIa, FcγRIIIb) and a dedicated FcRn adapter for pH-dependent IgG recycling kinetics.

Strategy: pre-conjugated adapter, then antibody analyte

Each adapter ships as a 110 µL stock at 2.6 µM in PBST — enough for 10 immobilisations at the recommended 200 nM working concentration (130 µL injection volume). The Fc receptor is supplied already-conjugated to a chosen oligonucleotide sequence (Seq ID 00–16), so multiple receptors with distinct Seq IDs can be loaded onto separate ridge positions in a single hybridisation step.

  1. FcγR / FcRn–oligo hybridisation. Flow the adapter (or a multiplex mix) at 200 nM in PBST over the Oligo|PEG chip for 5 min. The oligo hybridises to the matching ridge sequence, loading the Fc receptor on the surface.
  2. Antibody analyte injection. Run the antibody concentration series. Each receptor spot returns an independent sensorgram; for FcRn, run the binding phase at pH 6.0 and dissociate with pH 7.4 buffer.
  3. Regeneration. Strip the DNA duplex with 3 M GuHCl + 125 mM NaOH (400 µL/min, 0.5 min) to reload fresh receptor for the next antibody.

For non-standard or engineered Fc constructs not in the catalogue, conjugate the receptor off-chip with the AminoLink kit and load it the same way.

Adapter selection

AdapterReceptors coveredBest forWebshop
Oligo Adapter Fc gamma receptors (FcγRs)FcγRI (CD64), FcγRIIa (CD32a), FcγRIIb (CD32b), FcγRIIIa (CD16a), FcγRIIIb (CD16b) — all human; one Seq ID per receptor orderedFc-effector profiling across IgG subclasses; multiplex panels for engineered Fc variantsCHF 195 / 110 µL stock
Oligo Adapter FcRnHuman FcRn (Neonatal Fc Receptor)Antibody PK / half-life prediction; pH-dependent IgG recycling kinetics (binding pH 6.0, release pH 7.4)CHF 195 / 110 µL stock

Both adapters share the chip format and can be combined in a single multiplex hybridisation when their Seq IDs differ — profile FcγR binding and FcRn recycling for the same antibody in one experiment.

Protocol

Step 1 — Adapter hybridisation

ParameterValue
Adapter stock110 µL at 2.6 µM in PBST (10 immobilisations per vial)
Working dilution200 nM in PBST, 130 µL per injection
Flow rate / contact time10 µL/min, 5 min
Chip typeOligo|PEG (8×8 default; 6×9 on request)
MultiplexMix multiple adapters (different Seq IDs) at 200 nM each to load several receptors in one injection

Step 2a — Antibody analyte (FcγR adapter, pH 7.4)

ParameterValue
BufferPBST, pH 7.4
SCK association concentration range0.1–100 nM (high-affinity FcγRI), 1–1000 nM (low-affinity FcγRII / III)
SCK association flow rate / contact time30 µL/min (6-ch) / 60 µL/min (3–4-ch) / 100 µL/min (1-ch), 2 min
SCK dissociationPBST, 200 µL/min, 2 min (extend for slow off-rates)

Step 2b — Antibody analyte (FcRn adapter, pH 6.0)

ParameterValue
Binding buffer50 mM phosphate / 150 mM NaCl / 0.05 % Tween-20, adjusted to pH 6.0
SCK association concentration range10–1000 nM (FcRn affinity is moderate even at pH 6.0)
SCK association flow rate / contact time30 µL/min (6-ch) / 60 µL/min (3–4-ch) / 100 µL/min (1-ch), 2 min
SCK dissociationPBST pH 7.4, 200 µL/min, 2 min — IgG releases rapidly at neutral pH

Step 3 — Regeneration

ParameterValue
Regeneration solution3 M guanidinium-HCl + 125 mM NaOH
Flow rate / contact time400 µL/min, 0.5 min
After stripRe-hybridise the FcγR / FcRn adapter for the next antibody

Optimal parameters at a glance

ParameterFcγR adapterFcRn adapter
Stock110 µL at 2.6 µM in PBST (10 immobilisations per vial)
Working concentration200 nM, 130 µL injection
Hybridisation10 µL/min, 5 min, Oligo|PEG chip
Antibody analyte concentration0.1–100 nM (FcγRI) / 1–1000 nM (FcγRII–III)10–1000 nM
Binding buffer pH7.46.0 (binding) / 7.4 (release)
SCK association30 µL/min (6-ch) / 60 µL/min (3–4-ch) / 100 µL/min (1-ch), 2 min
SCK dissociation200 µL/min, 2 min (PBST or pH 7.4 release)
Regeneration3 M GuHCl + 125 mM NaOH, 400 µL/min, 0.5 min

Practical notes

See also: immobilisation method comparison table.