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Table 2 Ruminant and herbivorous species (bovine, sheep, horse and rabbit) intestinal organoids

From: Intestinal organoids in farm animals

Species Segment Application Crypt isolation Growth medium Reference
Bovine Jejunum S. Typhimurium and T. gondii infection 0.8 mM EDTA, 4 °C, 30 min 50% IntestiCult™ (STEMCELL Technologies, Vancouver, Canada) medium, 50% Wnt3a CM, 1 μg/mL rh R-spondin, 100 ng/mL rm Noggin, 100 ng/mL rm EGF, 1.5 μM CHIR99021, 5 μM Y27632, 5 μM SB202190, 250 nM A8301 [20]
  Ileum Model development 1% FBS, 75 U/mL collagenase 1-A, 20 µg/mL dispase I, 37 °C, 40 min IntestiCult™ (mouse) (STEMCELL Technologies, Vancouver, Canada), Y27632, LY2157299, SB202190 [22]
   E. coli infection 1% FBS, 75 U/mL collagenase 1-A, 20 µg/mL dispase I, 37 °C, 40 min IntestiCult™ (mouse) (STEMCELL Technologies, Vancouver, Canada), 10 µM Y27632, 500 nM LY2157299, 10 µM SB202190 [47]
   Rotavirus infection Not described DMEM/F12, 10 mM HEPES, GlutaMAX, 50 ng/mL EGF, 10% Noggin CM, 20% R-spondin CM, 50% Wnt3a CM, 10 mM nicotinamide, 10 nM gastrin I, 500 nM A-83–01, 10 μM SB202190, B27, N2, 1 mM NAC [24]
   Model development Collagenase, 37 °C, 1.5 h 50% (DMEM + 10% FBS), 50% L-WRN CM, 10 µM SB431542, 10 µM Y27632 [14]
  Colon 2D monolayer, in-plate cryoconservation Not described DMEM/F12, 1% BSA, 2 mM Glutamine, N2, B27, 10 mM HEPES, 50% Wnt3a-CM, 1 µg/mL rh R-spondin, 100 ng/mL rh Noggin, 50 ng/mL rh EGF, 500 nM A-83–01, 10 μM SB202190, 10 nM [Leu]15-gastrin-1, 10 mM nicotinamide, 1 mM NAC, 5 μM CHIR99021, 10 μM Y-27623, 50 nm Prostaglandin E2, 2 mM sodium acetate [23]
Sheep Ileum Model development Collagenase, 37 °C, 1.5 h 50% (DMEM + 10% FBS), 50% L-WRN CM, 10 µM SB431542, 10 µM Y27632 [14]
Horse Ileum Model development Collagenase, 37 °C, 1.5 h 50% (DMEM + 10% FBS), 50% L-WRN CM, 10 µM SB431542, 10 µM Y27632 [14]
  Jejunum Model development 30 mM EDTA, 10 mM Y27632, 1 mM DTT, 4 °C, 30 min + 30 mM EDTA, 10 mM Y27632, 37 °C, 10 min DMEM/F12, N-2, B27, GlutaMAX, 1 mM Hepes, 100 ng/mL hr noggin, 500 ng/mL rh R-spondin, 50 ng/mL hr EGF, 100 ng/mL, hr Wnt3a, 10 mM Y27632, 10 mM SB202190, 500 nM LY215799, 2,5 µM CHIR99021 [27]
Rabbit Caecum Model development 9 mM EDTA, 3 mM DTT, 10 µM Y27632, room temperature, 30 min Medium 1: DMEM, 10% FBS, 1 mM HEPES, 0.5 mM NAC, 10 µM CHIR99021, 10 µM Y27632,10 µM SB431542, 0.2 µM LDN193189
Medium 2: 50% (DMEM + 10% FBS), 50% L-WRN conditioned medium, 10 µM Y27632, 10 µM SB431542
[16]
  1. CM: conditioned medium, NAC: N-acetyl cysteine, L-WRN: L cell line engineered to secrete Wnt3a, R-spondin 3 and Noggin, rm: recombinant mouse, rh: recombinant human, FBS: fetal bovine serum, EGF: epidermal growth factor. Antibiotics and antifungals added in the growth medium are not presented.