Recent advances in flow cytometry instrumentation and fluorochrome chemistries have greatly increased fluorescent conjugated antibody combinations that can be used reliably and easily in routine experiments. The Cytek Aurora flow cytometer was first released with three excitation lasers (405, 488, and 640 nm) and incorporated the latest Avalanche Photodiode (APD) technology, demonstrating significant improvement in sensitivity for fluorescent emission signals longer than 800 nm.
However, there are limited commercially available fluorochromes capable of excitation with peak emission signals beyond 800 nm. To address this gap, we engineered six new fluorochromes: PE-750, PE-800, PE-830 for the 488 nm laser and APC-750, APC-800, APC-830 for the 640 nm laser.
Utilizing the principal of fluorescence resonance energy transfer (FRET), these novel structures were created by covalently linking a protein donor dye with an organic small molecule acceptor dye. Additionally, each of these fluorochrome conjugates were shown to be compatible with fixation/permeabilization buffer reagents, and demonstrated acceptable brightness and stability when conjugated to antigen-specific monoclonal antibodies. These six novel fluorochrome reagents can increase the numbers of fluorochromes that can be used on a spectral flow cytometer.
Immune Checkpoint LAG3 and Its Ligand FGL1 in Cancer
LAG3 is the most promising immune checkpoint next to PD-1 and CTLA-4. High LAG3 and FGL1 expression boosts tumor growth by inhibiting the immune microenvironment. This review comprises www.joplink.net/monoclonal-antibodies/ four sections presenting the structure/expression, interaction, biological effects, and clinical application of LAG3/FGL1. D1 and D2 of LAG3 and FD of FGL1 are the LAG3-FGL1 interaction domains. LAG3 accumulates on the surface of lymphocytes in various tumors, but is also found in the cytoplasm in non-small cell lung cancer (NSCLC) cells. FGL1 is found in the cytoplasm in NSCLC cells and on the surface of breast cancer cells.
The LAG3-FGL1 interaction mechanism remains unclear, and the intracellular signals require elucidation. LAG3/FGL1 activity is associated with immune cell infiltration, proliferation, and secretion. Cytokine production is enhanced when LAG3/FGL1 are co-expressed with PD-1. IMP321 and relatlimab are promising monoclonal antibodies targeting LAG3 in melanoma.
The clinical use of anti-FGL1 antibodies has not been reported. Finally, high FGL1 and LAG3 expression induces EGFR-TKI and gefitinib resistance, and anti-PD-1 therapy resistance, respectively. We present a comprehensive overview of the role of LAG3/FGL1 in cancer, suggesting novel anti-tumor therapy strategies.
Pharmacokinetics, Tolerability, Safety, and Immunogenicity of LY01008 and Bevacizumab (Avastin) in Healthy Chinese Subjects
Background and objective: LY01008 had been identified as being highly similar to the bevacizumab reference product in the pharmacy and pharmacology terms. The primary objective of this study was to compare the pharmacokinetic characteristics of the biosimilar candidate LY01008 with that of the bevacizumab (Avastin®) reference product after a single intravenous infusion in healthy Chinese adults. The secondary objective was to compare the safety and immunogenicity of LY01008 with those of bevacizumab.
Methods: In this double-blind, parallel-group, phase I study, 102 male subjects aged 18-45 years were randomized 1:1 to receive a single intravenous infusion of 3 mg/kg LY01008 or bevacizumab. Before the pivotal section, 12 healthy male subjects receiving a single intravenous (IV) infusion of 0.5 mg/kg or 1.5 mg/kg LY01008 were screened to verify the safety and tolerability of LY01008. Primary endpoints included the area under the concentration-time curve (AUC) from time zero to the last quantifiable time point (AUC0-t), AUC from time zero to the infinity time (AUC0-inf), and maximum plasma concentration (Cmax).
Results: The geometric mean ratios (GMRs) (90% confidence intervals, CIs) of AUC0-t, AUC0-inf, and Cmax of LY01008 to bevacizumab were 87.62% (82.91%, 92.61%), 87.27% (82.46%, 92.35%), and 96.45% (91.37%, 101.81%), respectively, in the pivotal section, which were within the prespecified equivalence margin of 80.00-125.00%. LY01008 and bevacizumab administered as a single 3 mg/kg intravenous dose were comparably well tolerated. No new or unexpected adverse events were observed. Nine subjects had antidrug antibodies (ADAs) (5 in the LY01008 group and 4 in the bevacizumab group) after dosing. No neutralizing antibody (Nab) was detected.
Meningoencephalitis caused by Fusarium proliferatum: an unusual case
Meningoencephalitis can be a diagnostic dilemma and one of its etiology are infectious causes including fungal agents. Fusarium species have attracted much attention as one of the invasive fungal infections. Major clinical manifestations of infections due to Fusarium spp. are broad such as keratitis, endophthalmitis, sino-pulmonary and central nervous system (CNS) infections. However, CNS fusariosis is rare and often happens due to hematogenous dissemination from other sites. Herein, we describe an unusual case of meningoencephalitis caused by Fusarium proliferatum, in a patient with rheumatoid arthritis.
Cis p-tau underlies vascular contribution to cognitive impairment and dementia, but is effectively targeted by immunotherapy
Background: Compelling evidence supports vascular contributions to cognitive impairment and dementia (VCID) including Alzheimer’s disease (AD), but the underlying pathogenic mechanisms and treatments are not fully understood. Cis P-tau is an early driver of neurodegeneration resulting from traumatic brain injury, but its role in VCID remains unclear.
Method: We first evaluated cis P-tau induction in VCID patients with no obvious AD pathology, and in mice after bilateral carotid artery stenosis (BCAS), a surgery modeling key aspects of clinical VCID. We subsequently tested if cis P-tau elimination using cis P-tau monoclonal antibody (cis mAb) reversed the VCID relevant pathology. To interrogate the upstream mechanism, we used the transgenic mice perturbing the cis P-tau counteracting isomerase Pin1 and Pin1’s inhibitory kinase DAPK1, and asked if perturbation of Pin1 and DAPK1 alters cis P-tau and VCID pathology. To investigate the mechanism underlying cis P-tau pathology, we dissected the BCAS mice cortical cell-type specific transcriptome with or without the cis mAb treatment. Finally, to test if cis P-tau is sufficient to induce the pathology, we stereotaxically injected the cis P-tau purified from the injury mice brain into injury-free mice cortex and evaluate their pathological and behavioral outcome.
Result: We surprisingly find robust and early cis P-tau despite no tau tangles in VCID patients and in mice modeling key aspects of clinical VCID, likely due to the inhibition of Pin1 by DAPK1. Elimination of cis P-tau in VCID mice using cis-targeted immunotherapy, brain-specific Pin1 overexpression or DAPK1 knockout effectively rescues VCID-like neurodegeneration and cognitive impairment in executive function. Furthermore, single-cell RNA-sequencing reveals that young VCID mice display diverse cortical cell-type specific transcriptomic changes resembling old AD patients, and the vast majority of these global changes are remarkably recovered by cis-targeted immunotherapy. Finally, purified soluble cis P-tau is sufficient to induce progressive neurodegeneration and brain dysfunction by causing axonopathy and conserved transcriptomic signature found at VCID mice and AD human patients with early pathologies.
Non-denaturing affinity purification of soluble oligomeric Aβ from human brain using a novel calcium-sensitive monoclonal antibody
Background: The soluble fraction of human AD brain extracts contains the most bioactive oligomeric forms of Aβ (oAβ), which are low in abundance. Few methods exist to enrich for oAβ from soluble brain extracts without denaturing them. Purification of natural, human oAβ would allow further structural and biochemical study of this truly disease-relevant species.
Method: A panel of anti-oAβ monoclonal antibodies was developed by immunizing Trianni mice with synthetic Aβ aggregates. Clones were screened for oAβ selectivity and protection against human oAβ-induced injury in an in vitro neuritic integrity assay and in mouse hippocampal slice LTP recordings. Four antibodies with selectivity for oAβ over monomeric Aβ and protective effects were chosen for further study.
Quantitative immunoprecipitations (IP followed by guanidine denaturation and monomer-specific ELISA) were performed on soluble extracts obtained by soaking cortical fragments bits in TBS to enrich for bioactive diffusible oAβ species. One antibody, B24, could IP oAβ only from non-dialyzed extracts. Iterative IPs were used to identify calcium as the soluble, dialyzable factor required for B24 binding to oAβ. Differential Scanning Fluorimetry (nano-DSF), Octet biolayer interferometry (BLI), and immunohistochemistry were further used to characterize B24 calcium-dependence and reversibility of binding to oAβ.
Result: B24 showed complete dependence on calcium at low millimolar levels for binding to soluble human oAβ, plaques, and synthetic Aβ protofibrils. Calcium induced a conformational change in B24. Binding of human soluble brain extracts to B24, washing, and elution with EGTA resulted in enrichment of oAβ. The recovered oAβ accounted for 20-40% of the initial input oAβ, accompanied by >400-fold reduction in total protein content. The recovered oAβ required guanidine hydrochloride for denaturation and detection by Aβ monomer-specific ELISA, implying that the recovered oAβ had not been denatured during purification. Calcium did not affect the size distribution of oAβ from human brain extracts by size exclusion chromatography.
YWHAZ Monoclonal Antibody |
|
| 0.05mL | 220 EUR |
YWHAZ Monoclonal Antibody |
|
| 0.1mL | 300 EUR |
YWHAZ Monoclonal Antibody |
|
| 5x0.1mL | 1350 EUR |
Monoclonal YWHAZ Antibody |
|
| 0.1ml | 580.8 EUR |
YWHAZ Mouse Monoclonal Antibody |
|
| 100 μl | 275 EUR |
Monoclonal YWHAZ Antibody, Clone: 1314CT423.108.153.173.140 |
|
| 0.1ml | 580.8 EUR |
Anti-14-3-3 zeta/delta/YWHAZ Antibody (Monoclonal, 6G5) |
|
| 0.1mg | 450 EUR |
Anti-14-3-3 zeta/delta/YWHAZ Antibody (Monoclonal, 6G5) |
|
| 5x0.1mg | 1870 EUR |
Anti-14-3-3 zeta/delta/YWHAZ Antibody (Monoclonal, 6H7) |
|
| 0.1mg | 450 EUR |
Anti-14-3-3 zeta/delta/YWHAZ Antibody (Monoclonal, 6H7) |
|
| 5x0.1mg | 1870 EUR |