mouse monoclonal α islet1 2 antibody  (Developmental Studies Hybridoma Bank)

Journal: Islets

Article Title: LDB1-mediated transcriptional complexes are sensitive to islet stress

doi: 10.1080/19382014.2021.2016028

Figure Lengend Snippet: Glucose levels modulate β-cell enriched mRNAs, including Isl1 . A-H. Relative mRNA quantification of various β-cell factors ( Insulin I, Insulin II, MafA, Pdx1, Nkx6.1, SSBP3, Isl1 , and Ldb1 respectively) from primary mouse islets after overnight culturing in media with increasing glucose concentrations (4 mM, 7.8 mM, 15.6 mM, and 31 mM). Low glucose (4 mM) was set as onefold, and 36B4 was used as the housekeeping gene; n = 3 for each treatment group. I. Top: Western blotting of Ins-1 cell extracts for LDB1, ISL1, or MAFA after 4 hr or overnight treatment with similar glucose concentrations as above. Actin was included as a loading control. Bottom: Densitometry quantification of 4 h or overnight treated LDB1, ISL1, or MAFA Western blot protein levels, normalized to Actin. *, P < .05; **, P < .01; ***, P < .001; ****, P < .0001 based on one-way ANOVA.

Article Snippet: After stimuli treatment for 4 hr in low (2.5 mM) or high (25 mM) glucose, the cells were formaldehyde fixed and then incubated with rabbit α-LDB1 (kind gift from Dr. Paul Love, NIH) and mouse α-ISL1 antibodies (39.4D5-c, Developmental Studies Hybridoma Bank) prior to PLA procedure.

Techniques: Western Blot, Control

Journal: Islets

Article Title: LDB1-mediated transcriptional complexes are sensitive to islet stress

doi: 10.1080/19382014.2021.2016028

Figure Lengend Snippet: Palmitate treatment downregulates LDB1 and SSBP3 co-regulator levels. A-H. Relative mRNA quantification of various β-cell factors ( Insulin I, Insulin II, MafA, Pdx1, Nkx6.1, SSBP3, Isl1 , and Ldb1 , respectively) from primary mouse islets after overnight culturing in media with control BSA (set as 1-fold) or palmitic acid (PA, 0.5 mM). 36B4 was used as the housekeeping gene; n = 3 for each treatment group. I . Left: Western blot of LDB1, MAFA, and ISL1 in Ins-1 cell extracts after overnight treatment with BSA control or 0.5 mM PA. Actin was included as loading control. Right: Densitometry quantification of LDB1, ISL1, or MAFA Western blot protein levels, normalized to Actin. *, P < .05; ***, P < .001; ****, P < .0001, based on Student’s t-test.

Article Snippet: After stimuli treatment for 4 hr in low (2.5 mM) or high (25 mM) glucose, the cells were formaldehyde fixed and then incubated with rabbit α-LDB1 (kind gift from Dr. Paul Love, NIH) and mouse α-ISL1 antibodies (39.4D5-c, Developmental Studies Hybridoma Bank) prior to PLA procedure.

Techniques: Control, Western Blot

Journal: Islets

Article Title: LDB1-mediated transcriptional complexes are sensitive to islet stress

doi: 10.1080/19382014.2021.2016028

Figure Lengend Snippet: Critical β-cell factors are reduced upon cytokine treatment in mouse and human islets. A-F. Relative mRNA quantification of various β-cell transcriptional regulator genes ( MafA, Pdx1, Nkx6.1, Ldb1, Isl1 , and SSBP3 , respectively) from primary mouse islets after 4 h culturing with single cytokines (Tnfα, Ifnγ, IL-1β) or a cocktail of each, as compared to PBS vehicle controls (set to 1-fold). 36B4 was used as the housekeeping gene; n = 3–4 for each treatment group. G. Left: LDB1, MAFA, NKX6.1, and ISL1 Western blot with Ins-1 cell extracts after 4 h or overnight treatment with single cytokines, or a cocktail, as compared to control. Actin was included as loading control. Right: Densitometry quantification of 4 h or overnight treated LDB1, MAFA, NKX6.1, or ISL1 Western blot protein levels, normalized to Actin. H. Human islets were treated with a DMSO vehicle or a cytokine cocktail, , then mRNA measured. 18S was used as the housekeeping gene; n = 3. I. Quantification of β-cell mRNAs from primary mouse islets after siRNA-mediated Ldb1 knockdown using Gapdh as the housekeeping gene; n = 3. *, P < .05; **, P < .01; ***, P < .001; ****, P < .0001 based on one-way ANOVA or Student’s t-test.

Article Snippet: After stimuli treatment for 4 hr in low (2.5 mM) or high (25 mM) glucose, the cells were formaldehyde fixed and then incubated with rabbit α-LDB1 (kind gift from Dr. Paul Love, NIH) and mouse α-ISL1 antibodies (39.4D5-c, Developmental Studies Hybridoma Bank) prior to PLA procedure.

Techniques: Western Blot, Control, Knockdown

Journal: Islets

Article Title: LDB1-mediated transcriptional complexes are sensitive to islet stress

doi: 10.1080/19382014.2021.2016028

Figure Lengend Snippet: Glucose and cytokines regulate LDB1:ISL1 protein interactions in Min6 β-cells. A-C. PLA immunofluorescence (red focal spots) and DAPI nuclear signals (blue) in Min6 cells treated with (A) control low glucose, (B) high-glucose, and (C) TNFα/IL-1β/IFNγ cytokine cocktail. Insets are 200% magnified images to show greater detail, which are taken from the main field, denoted by dashed white boxes. D. Quantification of nuclei lacking at least 1–2 nuclear PLA signals (red focal spots) in Min6 cells in A-C. E-F. PLA immunofluorescence (red) and DAPI nuclear signals (blue) in Min6 cells treated with (E.) Palmitate control-BSA or (F.) palmitic acid (PA). G. Quantification of nuclei lacking at least 1–2 nuclear PLA signals (focal spots) in Min6 cells in E-F. n = 3; scale bar = 20 μm. ****, P < .0001 based on Student’s t-test.

Article Snippet: After stimuli treatment for 4 hr in low (2.5 mM) or high (25 mM) glucose, the cells were formaldehyde fixed and then incubated with rabbit α-LDB1 (kind gift from Dr. Paul Love, NIH) and mouse α-ISL1 antibodies (39.4D5-c, Developmental Studies Hybridoma Bank) prior to PLA procedure.

Techniques: Immunofluorescence, Control

Journal:

Article Title: Control of her1 expression during zebrafish somitogenesis by a Delta -dependent oscillator and an independent wave-front activity

doi:

Figure Lengend Snippet: aei/DeltaD is required for both neurogenesis and patterning of aei/DeltaD expression in the PSM. (A–C) Examination of Islet-1-expressing neurons (black/blue) in the trunk of aei embryos revealed a neuronal hyperplasia. Using myosin (brown) as a reference for somite position, Islet-1-expressing neurons anterior to the posterior border of somite 5 were counted in wild-type (A) and aei embryos (B,C). Wild-type embryos (n = 20) averaged 33 Islet-1-expressing neurons in this region, whereas aeiAG49 (n = 19) and aeiAR33 (n = 20) averaged 58 and 61 such neurons, respectively. All embryos are at about the 13-somite stage. Photos show dorsal views of the trunk region. (D–I) Expression of aei/DeltaD is mispatterned in each of the fss-type mutants. Note, in fss embryos (F), 10%–15% show some evidence of aei/DeltaD stripe formation. Most embryos, however, exhibit expression patterns as seen in aei/DeltaD and the other fss-type mutants. All embryos are at about the 15-somite stage. Photos show dorsal views of the tailbud and posterior trunk. In all panels, anterior is to the left and posterior to the right.

Article Snippet: For antibody stainings, embryos were fixed in 4% PFA, blocked, incubated with the α-Islet-1 antibody (39.4D5 monoclonal, 1:500 dilution; Developmental Studies Hybridoma Bank), washed, incubated with 2% biotinylated anti-mouse (Vector), washed, and stained according to standard procedures.

Techniques: Expressing