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No effects on tubule length or branch-point formation were seen on cells either irradiated with light alone, or treated with drug alone.
After 48 h, the cells were either irradiated with 4 Gy IR or were mock-irradiated.
After 24 h incubation, cells were either irradiated with doses of 2 Gy or 4 Gy gamma radiations using a Co source (Theratron 1000, Theratronics, Inc., Ottawa, Ontario, Canada), or exposed first to L-OHP (50 or 100 μg/ml) for 2 h and afterward gamma-irradiated.
Bovine mammary cells were then added together with 2×105 NIH 3T3 mouse fibroblasts previously either irradiated with 50 Gy 250 KvP X-rays or treated with 10 µg/ml mitomycin C (Sigma-Aldrich, Milan, Italy) for 2 hours in human EpiCult B medium supplemented with 5% FBS, 10−6 M hydrocortisone (Sigma-Aldrich), 100 U/ml penicillin and 100 µg/ml streptomycin.
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After isolation, the hepatocytes were either irradiated (8 Gy) with 6 mV Photons using Variance Clinac 600C accelerator [ 17] or irradiated and additionally exposed to 500 ng of recombinant interleukin (IL) 1 β, IL-6, or TNF- α (PeproTech, Rocky Hill, NJ) in 3 mL cell culture supernatants immediately before irradiation.
To study this possibility, we incubated the Anti-p277 line for 3 days with either irradiated APC and p277 peptide or with irradiated, activated A2b T cells.
Sera from mice primed with either irradiated sporozoites or a synthetic peptide, (T1BT*)(4)-P3C, and boosted with (NANP)(3 -Q11 showed significant increases in antibody titers and significant inhibition of sporozoite infection in TSN assays.
Figure 3 shows that the ability of TregKIF cells to regulate CD25−CD4+ cells was not impaired, when cells were stimulated with either irradiated B10 APC (Fig 3a) or CD3/CD28 expander beads (Fig 3b).
Longer incubation of either irradiated, or untreated, cells with actinomycin D (up to 3 h) led to loss of the INB (data not shown), suggesting that it is dependent on intact nucleolar structure and/or nucleolar transcription.
On the other hand, it seems unlikely that an enhanced direct interaction between stromal fibroblasts and tumor cells could account for the earlier tumor establishment in D849N-mutant mice, since there was no difference in proliferation of B16 melanoma cells co-cultured with either irradiated wild type or irradiated D849N-mutant mouse embryonic fibroblasts in vitro (data not shown).
Total RNA was extracted from transfected cells (either irradiated or non-irradiated and with or without transfection) using TRIZOL reagent (Invitrogen, CA) as per standard protocol.
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