Adhesion of Day 3 P-Ir Thymocytes to Thymic Epithelial Cell Monolayers in the Presence of MoAb Directed to FN Receptors or Peptides Mimicking FN or LN Moieties
| Addition . | Amount . | Concentration (mmol/L) . | Relative Binding . | |
|---|---|---|---|---|
| Exp 1 . | Exp 2 . | |||
| No | — | — | 100.0* | 100.0* |
| Rat Ig | 20 μg/mL | — | 99.6 | 89.2 |
| Anti-α4 chain antibody | 20 μg/mL | — | 43.7 | 37.2 |
| Anti-α5 chain antibody | 20 μg/mL | — | 100.7 | 93.2 |
| FN 1-25 IIICS (LDV)†,‡ | 1 mg/mL | 0.37 | 23.5 | 30.0 |
| FN 90-109 IIICS (REDV)†,‡ | 1 mg/mL | 0.42 | 77.8 | 79.6 |
| FN GRGDS‡ | 1 mg/mL | 2.04 | 81.9 | 83.6 |
| LN 925-933 (YIGSR)‡ | 1 mg/mL | 1.03 | 102.4 | 87.6 |
| Addition . | Amount . | Concentration (mmol/L) . | Relative Binding . | |
|---|---|---|---|---|
| Exp 1 . | Exp 2 . | |||
| No | — | — | 100.0* | 100.0* |
| Rat Ig | 20 μg/mL | — | 99.6 | 89.2 |
| Anti-α4 chain antibody | 20 μg/mL | — | 43.7 | 37.2 |
| Anti-α5 chain antibody | 20 μg/mL | — | 100.7 | 93.2 |
| FN 1-25 IIICS (LDV)†,‡ | 1 mg/mL | 0.37 | 23.5 | 30.0 |
| FN 90-109 IIICS (REDV)†,‡ | 1 mg/mL | 0.42 | 77.8 | 79.6 |
| FN GRGDS‡ | 1 mg/mL | 2.04 | 81.9 | 83.6 |
| LN 925-933 (YIGSR)‡ | 1 mg/mL | 1.03 | 102.4 | 87.6 |
Thymocytes (1.5 × 107 in 1 mL FCS-free medium) were pre-incubated in the presence of MoAb or peptides for 30 minutes on ice bath and then added to 2BH4 monolayers. After an additional incubation of 1 hour at 37°C, nonadherent thymocytes were removed by aspiration followed by three washes with warm medium. Adherent thymocytes were recovered in cold medium with FCS after beating the culture flasks and pipette flushing.
100% represents 7.7% and 4.3% of total thymocytes used in Exp 1 and Exp 2, respectively.
IIICS means the alternatively spliced type III connecting segment from FN.
In bold are shown the amino acid sequences critical for integrin binding (LDV or REDV for α4β1, RGD for α5β1, and YIGSR for the 67-kD LN receptor).