- PA5-35136 - Invitrogen Antibodies SLC11A2 antibody

Lv H, Zhen C, Liu J, Shang P
Oxidative medicine and cellular longevity 2020;2020:5021983

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of SLC11A2 in mouse cerebellum tissue lysate (35 µg/lane) using a SLC11A2 polyclonal antibody (Product # PA5-35136).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis was performed on membrane enriched extracts (30 µg lysate) of HepG2 (Lane 1), SH-SY5Y (Lane 2), Caco-2 (Lane 3), COLO 205 (Lane 4), HCT 116 (Lane 5), HT29 (Lane 6), HeLa (Lane 7) and PC-3 (Lane 8). The blot was probed with Anti-SLC11A2 Polyclonal Antibody (Product # PA5-35136, 1:2000 dilution) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25 µg/ml, 1:4000 dilution). A 50 kDa band corresponding to SLC11A2 was observed across the cell lines tested.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent analysis of SLC11A2 in HepG2 cells using a SLC11A2 polyclonal antibody (Product # PA5-35136) followed by detection using a fluorescent conjugated secondary antibody (green). Nuclei were stained with Dapi (blue).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of SLC11A2 was performed using 70% confluent log phase Hep G2 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with SLC11A2 Polyclonal Antibody (Product # PA5-35136) at 1:50 dilution in 0.1% BSA, incubated at 4 degree Celsius overnight and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing cytoplasmic localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 4 PEITC altered iron metabolism in human OS cells. (a) Total iron level in MNNG/HOS, U-2 OS, MG-63, and 143B cells after the indicated concentrations of PEITC treatment for 24 h by AAS. (b) The level of labile iron in MNNG/HOS, U-2 OS, MG-63, and 143B cells after the indicated concentrations of PEITC treatment for 24 h with Calcein-AM staining by flow cytometry analysis. (c) Protein expression levels of TfR1, DMT1, FTH1, FPN, and IRP2 in MNNG/HOS, U-2 OS, MG-63, and 143B cells treated with the indicated concentrations of PEITC for 20 h or 30 mu M PEITC for 4 h, 12 h, 24 h, and 48 h. (d) mRNA levels of TFRC , SLC11A2 , FTH1 , SLC40A1 , and IRP2 in MNNG/HOS, U-2 OS, MG-63, and 143B cells after PEITC treatment for 24 h. (e) Protein expression levels of NCOA4 in MNNG/HOS, U-2 OS, MG-63, and 143B cells treated with the indicated concentrations of PEITC for 20 h or 30 mu M PEITC for 4 h, 12 h, 24 h, and 48 h. All data were presented as the means +- SD ( n = 3). * P < 0.05, ** P < 0.01, and *** P < 0.001 versus control group.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 9 PEITC induced cell death via ROS generation in human OS cells. (a) EdU staining assay of MNNG/HOS, U-2 OS, MG-63, and 143B cells treated with 30 mu M PEITC in the presence or absence of NAC for 24 h. (b) Quantitative analysis of EdU staining in (a). (c) Protein expression levels of TfR1, DMT1, FTH1, and FPN in MNNG/HOS, U-2 OS, MG-63, and 143B cells treated with 30 mu M PEITC in the presence or absence of NAC for 24 h. (d) Protein expression levels of GPx4, LC3B, C-PARP, C-caspase3, Bcl2, and Cdc2 in MNNG/HOS, U-2 OS, MG-63, and 143B cells treated with 30 mu M PEITC in the presence or absence of NAC 24 h. (e) Phosphorylation levels of ERK, p38, and JNK in MNNG/HOS, U-2 OS, MG-63, and 143B cells treated with 30 mu M PEITC in the presence or absence of NAC for 24 h.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 10 PEITC suppressed OS growth in vivo . MNNG/HOS cells were injected subcutaneously into the right flank of the male BALB/c nude mice. One week after the OS xenograft mouse model was established, the mice were randomly divided into two groups and, respectively, administrated with 10% sesame seed oil and 30 mg/kg PEITC once daily for 24 consecutive days. (a) Body weight change of the two groups. (b) Volume change of OS tissues of the two groups. Data were calculated by the formula: volume = length x width 2 x 0.5. (c) Weight of OS tissues from the two groups. (d) H&E staining analysis of tumor tissues (200x and 400x). (e) The protein expression levels of TfR1, DMT1, FTH1, and FPN in tumor tissues. (f) The protein expression levels of LC3B, C-caspase3, GPx4, and Cdc2 in tumor tissues. (g) Phosphorylation levels of ERK, p38, and JNK in tumor tissues. All data were presented as the means +- SD ( n = 7). * P < 0.05, ** P < 0.01, and *** P < 0.001 versus control group.

PA5-35136