Journal: Bio-protocol

Article Title: Quantification of 20-Hydroxyecdysone From Drosophila Larvae at the Onset of Metamorphosis

doi: 10.21769/BioProtoc.5831

Figure Lengend Snippet: Preparation of different standard solutions (Std1–7 and B0) of 20E for ELISA using 1× assay buffer as diluent.

Article Snippet: ELISA plate shaker (Neuation, model: iSHAK TS4 NXT)

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Bio-protocol

Article Title: Quantification of 20-Hydroxyecdysone From Drosophila Larvae at the Onset of Metamorphosis

doi: 10.21769/BioProtoc.5831

Figure Lengend Snippet: Reagent and buffer volumes for competitive ELISA setup

Article Snippet: ELISA plate shaker (Neuation, model: iSHAK TS4 NXT)

Techniques: Competitive ELISA, Enzyme Immunoassay

Journal: Bio-protocol

Article Title: Quantification of 20-Hydroxyecdysone From Drosophila Larvae at the Onset of Metamorphosis

doi: 10.21769/BioProtoc.5831

Figure Lengend Snippet: The principle of 20E estimation by this method relies on competitive ELISA. The presence of unlabeled 20E in samples will compete with 20E-HRP for binding to anti-20E antibodies and enzymatic reaction-dependent color generation. (A) Blank represents anti-rabbit antibody-coated wells without any buffers or reagents (no binding, no reaction, and no color). (B) NSB indicates minimal or nonspecific background binding (nonspecific adherence of 20E-HRP to the substratum and thus background reactivity and color generation). (C) B0 represents the maximum binding of 20E-HRP in the absence of free 20E (no competition, maximum binding, maximum enzymatic activity, highest color generation). (D) Standard demonstrates the inverse relationship between 20E-HRP binding and the concentration of free 20E (available 20E amount-dependent competition, curtailed enzyme activity, reduced color generation).

Article Snippet: ELISA plate shaker (Neuation, model: iSHAK TS4 NXT)

Techniques: Competitive ELISA, Binding Assay, Nonspecific Binding, Saturation Binding, Activity Assay, Concentration Assay

Journal: bioRxiv

Article Title: Symmetric Dimethylarginine Modification Drives Shared and Divergent Antigen Reactivity in Lupus

doi: 10.64898/2026.09.22.753525

Figure Lengend Snippet: (A) Sera from MRL lpr mice (n=12, bled weekly from 8 to 13 weeks of age) were analyzed by ELISA using synthetic 10×RG peptides: native sequence (GRGRGRGRGRGRGRGRGRGR) vs. SDMA-modified (lowercase ‘r’ denotes SDMA; GrGrGrGRGRGrGrGrGRGR). All 12 mice developed anti SDMA antibody titers above the corresponding native controls, and the majority (9/12) showed a gradual increase in anti-SDMA titers over time. (B) Immunoblotting of HEp-2 lysates revealed progressive changes in band patterns from 10 weeks onward. To identify SDMA specific reactive proteins, cells were pretreated with the methyltransferase inhibitor EPZ015666 (EPZ) before lysis. A prominent ∼15 kDa band (arrowhead) was markedly reduced by EPZ pretreatment, supporting its recognition by anti SDMA antibodies. (C) Sera from three 12 week old MRL lpr littermates (Lpr 1, 2, 3) each showed distinct band patterns against the cell lysates. EPZ pretreatment of the cells reduced the intensity of several bands in all three mice. (D) Workflow for isolating hybridoma clones from individual plasma cells to obtain monoclonal antibodies (mAbs) specific for SDMA. MRL lpr mice were not immunized with any antigen; thus, the resulting mAbs represent naturally arising autoantibodies. (E) Two antibody clones were tested against the native versus SDMA peptide array shown in Figurè3. Clone P4A2F4 reacted extensively with most SDMA containing peptides, whereas clone P2C1C10 showed no reactivity. (F) The two mAbs produced different band patterns on cell lysates. P4A2F4 recognized bands that were reduced in EPZ pretreated lysates, consistent with SDMA specificity. (G) After implantation of the SDMA-reactive hybridoma clone into the peritoneal cavity of healthy BALB/c mice, the kidneys showed extensive IgG deposition, reminiscent of human lupus nephritis.

Article Snippet: High-binding ELISA plates (Greiner Bio-One, 655081) were coated with 250 ng/well of streptavidin (Leinco Technologies, S203) overnight at 4°C.

Techniques: Enzyme-linked Immunosorbent Assay, Sequencing, Modification, Western Blot, Lysis, Clone Assay, Clinical Proteomics, Bioprocessing, Autoantibodies, Peptide Microarray, Produced