Anti vegf

Western blot analyses were performed as follows. Protein samples obtained from brain or placenta tissue homogenates were separated on 8%–12% sodium dodecyl sulfate‐polyacrylamide gels, transferred to Immobilon‐P membranes (Millipore), and probed overnight at 4°C with primary antibodies including anti‐BDNF (1:1000; GTX132621), anti‐GDNF (1:1000; GTX17447), and anti‐VEGF (1:1000; GTX102643) from GeneTex (Irvine, CA, USA); anti‐pCaMKII (1:1000; #12716), anti‐total CaMKII (1:1000; #11945), anti‐pAkt S473 (1:1000; #9271), anti‐pAktT308 (1:1000; #4056), anti‐total Akt (1:1000; #9272), anti‐pERK (1:1000; #9101), and anti‐total ERK (1:1000; #9102) from Cell Signaling; anti‐PlGF (1:1000; ab196666), anti‐glucose transporter (GLUT) 3 (ab41525), anti‐GLUT1 (ab115730), anti‐SLC38A1 (ab134268), anti‐SNAT2 (ab90677), and anti‐SLC38A4 (ab58785) from Abcam; and anti‐β‐actin (1:10000; A5441, Sigma‐Aldrich). Afterward, the membranes were washed and incubated with anti‐rabbit or anti‐mouse horseradish peroxidase‐linked secondary antibodies (1:5000; Santa Cruz Biotechnology, Santa Cruz, CA, USA) at 4°C for 1 h. The relative intensity of protein signals was normalized to the corresponding β‐actin intensity and was quantified by densitometric analysis with the use of ImageJ 1.50i software (National Institutes of Health, Bethesda, MD, USA).