2 7 dichlorodihydrofluorescein diacetate dcfh da

CCK‐8 kit and Annexin V‐FITC/PI Apoptosis Detection kit were purchased from Dojindo (Kumamoto, Japan). PI, Glutaraldehyde solution, 2′, 7′‐Dichlorodihydrofluorescein diacetate (DCFH‐DA) and N‐acetyl‐L‐cysteine (NAC) are from Sigma‐Aldrich (St Louis, MO, USA). 3‐Methyladenine (3‐MA), Bafilomycin A1 (Baf‐A1), Dorsomorphin (Compound C), SB202190, U0126 and SP600125 were obtained from Selleck (Houston, Texas, USA).4′,6‐diamidino‐2‐phenylindole (DAPI), Lyso‐Tracker Red and Lipid Peroxidation MDA Assay kit were purchased from Beyotime (Nantong, China).
Antibodies against Cyclin B1 (ET1608‐27), Cyclin A2 (M1511‐5), CDK2 (ET1602‐6), CDK1 (ET1605‐54), ATG5 (ET1611‐38), p38 (ET1602‐26), ERK1/2 (ET1601‐29), phospho‐ERK1/2(Thr202) (ET1610‐13), JNK1/2/3 (ET1601‐28), phospho‐JNK1/2/3(T183 + T183 + T221) (ET1609‐42), AMPK alpha 1 (ET1608‐40), phospho‐AMPK alpha 1 (S496) (ET1612‐72), HSPB1 (ET1701‐70), phospho‐HSPB1(S82) (ET1611‐16) and PARP (ET1608‐56) were purchased from HuaBio (Hangzhou, China). Antibodies against β‐actin (66009‐1‐lg), p53 (10442‐1‐AP), p21 (10355‐1‐AP), p62 (18420‐1‐AP), BAX (50599‐2‐1g) and caspase 3 (19677‐1‐AP) were obtained from Proteintech (Wuhan, China). The antibodies against phospho‐p38 (T180/Y182) (9211S) and LC3B (2775s) were purchased from Cell Signaling Technology (Beverly, MA, USA).

Blueberry powder and blackcurrant powder were purchased online (Viberi, Timaru, New Zealand). Oat bran was obtained from the local supermarket (Sun Valley, Christchurch, New Zealand). Pepsin (EC 3.4.23.1) pancreatin (EC 232-468-9), α-amyloglucosidase (EC 3.2.1.3), invertase (EC 3.2.1.26), 2,2′-Azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), 2,4,6-Tris(2-pyridyl)-s-triazine (TPTZ), 2-Diphenyl-1-picrylhydrazyl (DPPH), 3,5-Dinitrosalicylic acid (DNS, 98%, ACROS Organics™, Waltham, MA, USA ), Folin and Ciocalteu’s phenol reagent, 2,7 dichlorodihydrofluorescein-diacetate (DCFH-DA), gallic acid, rutin, and trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid) were all purchased from Sigma-Aldrich (St. Louis, MO, USA). Other chemicals in this study were of analytical grade.

Triplets for each bacterial strain tested were placed on a 96-well black microtiter plate for cell culture by adding an equal volume of colloidal platinum nanoparticle solution at the appropriate 2 × MIC, 1 × MIC, and ½ × MIC concentrations to the inoculum (0.5 McFarland units). The plate was incubated at 37 °C for 1, 3, 6, 12, and 24 h. After incubation, a peroxynitrite indicator, 2′-7′-dichlorodihydrofluorescein diacetate (DCFH-DA) (Sigma-Aldrich, Gillingham, UK) (final concentration: 5 µM) was added to the wells. In order to achieve better bonding of the dye to ROS, the light-protected plate was placed on an electric mixer for microtiter plates at medium speed (348/1 “ASSISTENT” Rocking shaker, CE type, Karl Hecht GmbH & Co., Sondheim vor der Rhön, Germany). Fluorescence intensity was measured at 485 nm excitation and 525 nm emission (Infinite 200 microplate reader, Tecan Group Ltd., Männedorf, Switzerland) [53 ,54 (link)].

Dulbeco's modified eagle medium (DMEM), fetal calf serum (FCS), penicillin, streptomycin, glutamate, phosphate buffered saline (PBS), crystal violet, 2′,7′-dichlorodihydrofluorescein-diacetate (DCFH-DA), 0,25% Triton-X-100, bovine serum albumin (BSA), cell dissociation solution (CDS), dodecyl-D-β- Maltosid, ammoniumbicarbonat (AB) were purchased from Sigma Aldrich (St. Louis, MO). L-alanyl-L-glutamin was purchased from Biochrom AG (Berlin, Germany). H₂O₂ and paraformaldehyde was obtained from Carl Roth GmbH (Karlsruhe, Germany), staurosporine was purchased from Calbiochem (San Diego, CA). Ethanol, acetonitril (ACN), trifluoroacetic acid (TFA) and formic acid were purchased from Merck (Darmstadt, Germany), wheat germ agglutinin from Invitrogen (Carlsbad, U.S.A.) and BCA Pierce Protein Assay kit and Dylight 649 was purchased from Fisher scientific (Waltham, MA.). Trypsin was from Promega (Mannheim, Germany), HPLC H₂O from Applichem (Darmstadt, Germany) and C-18 ZipTips was purchased from Millipore (Billerica, MA). The used abs were listed in table 1.