Bradford Assay:
Recombinant:
Article Title: The one-week automated genome-wide optical pooled screen
Article Snippet: Padlock probe gap-fill and ligation were performed in 1X Ampligase buffer (Lucigen A3210K) containing 50 nM dNTPs (NEB N0447L), 0.1 μM each padlock probe (Integrated DNA Technologies), 200 μg/mL recombinant albumin (NEB B9200S), 0.4 U/μL RNase H (Enzymatics Y9220L), 0.02 U/μL TaqIT polymerase (Enzymatics P7620L), and 0.5 U/μL Ampligase (Lucigen A3210K). .. Samples were incubated in gap-fill and ligation solution at 37 °C for 5 minutes followed by 45 °C for 90 minutes, then washed twice with PBS-T. Rolling circle amplification (RCA) was performed in 1X Phi29 buffer (Thermo Fisher Scientific EP0091) containing 250 μM dNTPs (NEB N0447L), 5% glycerol (MilliporeSigma G5516), 200 μg/mL recombinant albumin (NEB B9200S), and 1 U/μL Phi29 DNA polymerase (Thermo Fisher Scientific EP0091) for 12 hours at 30 °C. .. Samples were washed twice with PBS-T, then 1 μM sequencing primers (Integrated DNA Technologies) were hybridized in 2X SSC (Ambion AM9763) with 10% (v/v) formamide (Thermo Fisher Scientific 4650) for 30 minutes at room temperature and washed twice with PBS-T prior to in situ sequencing.
Article Title: The one-week automated genome-wide optical pooled screen
Article Snippet: Samples were washed three times with PBS-T and post-fixed in 3% formaldehyde (Electron Microscopy Sciences 15714) and 0.1% glutaraldehyde (Electron Microscopy Sciences 16120) for 30 minutes, followed by quenching for 1 minute in pH 8 0.2 M Tris-HCl (Thermo Fisher Scientific AM9855G) and three PBS-T washes. .. Reverse transcription was carried out in 1X RevertAid RT buffer (Thermo Fisher Scientific EP0452) containing 250 μM dNTPs (NEB N0447L), 1 μM biotinylated reverse-transcription primer (Integrated DNA Technologies), 200 μg/mL recombinant albumin (NEB B9200S), 0.8 U/μL RiboLock RNase inhibitor (Thermo Fisher Scientific EO0384), and 4.8 U/μL RevertAid H-minus Reverse Transcriptase (Thermo Fisher Scientific EP0452). ..
Article Title: H2A.Z facilitates Sox2-nucleosome interaction by promoting DNA and histone H3 tail mobility
Article Snippet: .. Reactions were set up in 10 mM Tris–HCl pH 7.5, 100 mM KCl, 1 mM DTT, 0.2 mg/ml recombinant albumin (NEB), 0.02% Tween-20, 8% glycerol, containing 100 nM Lin28B DNA or NCP. .. Binding reactions were set up with or without Sox2 HMG (0.1 μM or 1 μM) and Oct4 CtoS (0.3 μM or 3 μM), and incubated at 25°C for 45 min. DNaseI (NEB) was added at 0.5 U (or 0.125 U) for NCP and 0.1 U (or 0.025 U) for DNA and incubated for 5 min at 25°C, inactivated by 40 μl quench buffer [10 mM Tris–HCl pH 7.5, 50 mM EDTA, 2% SDS, 300 ng/μl glycogen (Invitrogen)], and heating at 75°C for 30 min, as previously described [ ].
Article Title: Dissecting the cellular architecture of breast cancer brain metastases reveals prognostically distinct immune landscapes.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Killing assay: Mouse monoclonal anti-CD28 (clone CD28.2) BioLegend Cat#302902 (RRID:AB_314304) ICB assay: Pembrolizumab Heidelberg University Hospital Pharmacy NA ICB assay: Tiragolumab Hycultec Cat# HY-P9986 (RRID:AB_3695514) Biological samples BCBM tissue specimens Biobank of the Department of Neurosurgery, Heidelberg University Hospital, Heidelberg, Germany N/A Blood samples of BCBM patients Biobank of the Department of Neurosurgery, Heidelberg University Hospital, Heidelberg, Germany N/A Chemicals, peptides, and recombinant proteins 2-Methylbutane Carl Roth Cat#3927.2 AMPure XP beads Beckman Coulter Cat#A63881 Biocoll solution Biochrom Cat#L6115 Bovine serum albumin (BSA), molecular biology grade New England Biolabs Cat#B9000S Brefeldin A Solution Biolgend Cat#420601 Buffer EB Qiagen Cat#19086 Calcium chloride (CaCl2) 1 M Hölzel Diagnostika Cat#103047-1 CellTiter-Blue assay Promega Cat#G8081 DAKO diluent, background reducing Agilent Dako Cat#S302283-2 DAPI (4’,6-Diamidin-2-phenylindol) Thermo Fisher Scientific Cat#D1306 Deoxyribonuclease I (DNAse I) Sigma-Aldrich Cat#DN25-100MG EGF Novoprotein Cat#C029 β-Estradiol Sigma-Aldrich Cat#E2758 bFGF Novoprotein Cat#C046 BIT admixture 100 PELOBiotech Cat#PB-SH-033-0000 DMEM/F-12, no phenol red Thermo Fisher Scientific Cat#21041025 DMEM, high glucose, GlutaMAX supplement Thermo Fisher Scientific Cat#61965059 Dulbecco’s Phosphate Buffered Saline (DPBS) Thermo Fisher Scientific Cat#14190-094 EDTA (Versen) 1% in PBS Genaxxon Bioscience Cat#C4263.0100 FCS (Fetal calf serum) Sigma-Aldrich Cat#S0615 Fixable Viability Dye eFluor 506 Thermo Fisher Scientific Cat#65-0866-14 Fixable Viability Dye eFluor 450 Thermo Fisher Scientific Cat#65-0863-14 GlutaMAX Supplement Thermo Fisher Scientific Cat#35050038 Glycerol Sigma-Aldrich Cat#1040950250 IgG from human serum Sigma-Aldrich Cat#I2511 IL-2, human, recombinant Stemcell Technologies Cat#78036.3 IncuCyte Caspase-3/7 Green Dye Sartorius Cat#4440 IncuCyte Cytolight Rapid Red Dye Sartorius Cat#4706 ImmunoCult Human CD3/CD28/CD2 T Cell Activator Stemcell Technologies Cat#10970 ImmunoCult-XF T Cell Expansion Medium Stemcell Technologies Cat#10981 Liberase DH Research Grade Merck Cat#05401089001 Methylene blue Conatex Cat#9993339 Magnesium chloride (MgCl2) 1 M Sigma Aldrich Cat#M1028 Mowiol 4-88 Carl Roth Cat#0713.2 (Continued on next page) ll OPEN ACCESSArticle Cancer Cell 44, 1–22.e1–e16, June 8, 2026 e3 .. REAGENT or RESOURCE SOURCE IDENTIFIER Nuclease-free water Thermo Fisher Scientific Cat#AM9937 Penicillin-Streptomycin (10.000 U/ml) Thermo Fisher Scientific Cat# 15140122 Propidium Iodide BioLegend Cat#421301 Sodium chloride (NaCl) 5 M Thermo Fisher Scientific Cat#AM9760G Staurosporine Hycultec Cat# HY-15141 Synth-a-Freeze cryopreservation medium Thermo Fisher Scientific Cat#A1254201 Tissue-Tek O.C.T. compound Science Services Cat#SA6255012 Tris (pH 7.5), 1 M Thermo Fisher Scientific Cat#15567027 Tween-20 Sigma-Aldrich Cat#P-7949 Critical commercial assays Chromium NEXT GEM Chip K Single Cell Kit 10x Genomics Cat#1000286 and Cat#1000287 Chromium Next GEM Single Cell 5’ Kit v2 - Chromium Next GEM Single Cell 5’ Gel Bead Kit v2 - Chromium Next GEM Single Cell 5ʹ GEM Kit v2 - Library Construction Kit 10x Genomics Cat#1000263 - Cat#1000264 and Cat#1000267 - Cat#1000244 and Cat#1000266 - Cat#1000190 and Cat#1000196 Tube, Dynabeads MyOne SILANE 10x Genomics Cat#2000048 Dual Index Kit TT Set A 10x Genomics Cat#1000215 EasySep Human CD3 Positive Selection Kit II Stemcell Technologies Cat#17851 Foxp3 / Transcription Factor Staining Buffer Set Thermo Fisher Scientific Cat# 00-5523-00 IC fixation buffer Thermo Fisher Scientific Cat# 00-8222-49 Infinium OncoArray-500K BeadChip Illumina Cat#WG-355-1001 LIVE/DEAD Cell Imaging Kit (488/570) Thermo Fisher Scientific Cat#R37601 Tumor Dissociation Kit, human Miltenyi Biotec Cat#130-095-929 Deposited data Bulk RNA-seq data, BCBM discovery cohort This paper German Human Genome-Phenome Archive (GHGA) as GHGAD28055171876192 Gene Expression Omnibus: GSE322943 Bulk RNA-seq data, BCBM CATCH cohort This paper German Human Genome-Phenome Archive (GHGA) as GHGAD51052817169855 Single nuclei RNA-seq data, BCBM This paper Zenodo at 10.5281/zenodo.17723784 German Human Genome-Phenome Archive (GHGA) as GHGAD48917471237543 Gene Expression Omnibus: GSE324453 Spatial transcriptomics data This paper Zenodo at https://doi.org/10.5281/zenodo.
Article Title: The one-week automated genome-wide optical pooled screen
Article Snippet: Samples were washed twice with PBS-T and incubated for 15 minutes with 20 μg/mL streptavidin (NEB N7021S) and 100 μg/mL recombinant albumin (NEB B9200S) in PBS, followed by two PBS-T washes and postfixation in 3% formaldehyde (Electron Microscopy Sciences 15714) and 0.1% glutaraldehyde (Electron Microscopy Sciences 16120) for 30 minutes. .. Padlock probe gap-fill and ligation were performed in 1X Ampligase buffer (Lucigen A3210K) containing 50 nM dNTPs (NEB N0447L), 0.1 μM each padlock probe (Integrated DNA Technologies), 200 μg/mL recombinant albumin (NEB B9200S), 0.4 U/μL RNase H (Enzymatics Y9220L), 0.02 U/μL TaqIT polymerase (Enzymatics P7620L), and 0.5 U/μL Ampligase (Lucigen A3210K). .. Samples were incubated in gap-fill and ligation solution at 37 °C for 5 minutes followed by 45 °C for 90 minutes, then washed twice with PBS-T. Rolling circle amplification (RCA) was performed in 1X Phi29 buffer (Thermo Fisher Scientific EP0091) containing 250 μM dNTPs (NEB N0447L), 5% glycerol (MilliporeSigma G5516), 200 μg/mL recombinant albumin (NEB B9200S), and 1 U/μL Phi29 DNA polymerase (Thermo Fisher Scientific EP0091) for 12 hours at 30 °C.
Lysis:
Incubation:Article Title: The one-week automated genome-wide optical pooled screen
Article Snippet: Padlock probe gap-fill and ligation were performed in 1X Ampligase buffer (Lucigen A3210K) containing 50 nM dNTPs (NEB N0447L), 0.1 μM each padlock probe (Integrated DNA Technologies), 200 μg/mL recombinant albumin (NEB B9200S), 0.4 U/μL RNase H (Enzymatics Y9220L), 0.02 U/μL TaqIT polymerase (Enzymatics P7620L), and 0.5 U/μL Ampligase (Lucigen A3210K). .. Samples were incubated in gap-fill and ligation solution at 37 °C for 5 minutes followed by 45 °C for 90 minutes, then washed twice with PBS-T. Rolling circle amplification (RCA) was performed in 1X Phi29 buffer (Thermo Fisher Scientific EP0091) containing 250 μM dNTPs (NEB N0447L), 5% glycerol (MilliporeSigma G5516), 200 μg/mL recombinant albumin (NEB B9200S), and 1 U/μL Phi29 DNA polymerase (Thermo Fisher Scientific EP0091) for 12 hours at 30 °C. .. Samples were washed twice with PBS-T, then 1 μM sequencing primers (Integrated DNA Technologies) were hybridized in 2X SSC (Ambion AM9763) with 10% (v/v) formamide (Thermo Fisher Scientific 4650) for 30 minutes at room temperature and washed twice with PBS-T prior to in situ sequencing.
Ligation:Article Title: The one-week automated genome-wide optical pooled screen
Article Snippet: Padlock probe gap-fill and ligation were performed in 1X Ampligase buffer (Lucigen A3210K) containing 50 nM dNTPs (NEB N0447L), 0.1 μM each padlock probe (Integrated DNA Technologies), 200 μg/mL recombinant albumin (NEB B9200S), 0.4 U/μL RNase H (Enzymatics Y9220L), 0.02 U/μL TaqIT polymerase (Enzymatics P7620L), and 0.5 U/μL Ampligase (Lucigen A3210K). .. Samples were incubated in gap-fill and ligation solution at 37 °C for 5 minutes followed by 45 °C for 90 minutes, then washed twice with PBS-T. Rolling circle amplification (RCA) was performed in 1X Phi29 buffer (Thermo Fisher Scientific EP0091) containing 250 μM dNTPs (NEB N0447L), 5% glycerol (MilliporeSigma G5516), 200 μg/mL recombinant albumin (NEB B9200S), and 1 U/μL Phi29 DNA polymerase (Thermo Fisher Scientific EP0091) for 12 hours at 30 °C. .. Samples were washed twice with PBS-T, then 1 μM sequencing primers (Integrated DNA Technologies) were hybridized in 2X SSC (Ambion AM9763) with 10% (v/v) formamide (Thermo Fisher Scientific 4650) for 30 minutes at room temperature and washed twice with PBS-T prior to in situ sequencing.
Article Title: The one-week automated genome-wide optical pooled screen
Article Snippet: Samples were washed twice with PBS-T and incubated for 15 minutes with 20 μg/mL streptavidin (NEB N7021S) and 100 μg/mL recombinant albumin (NEB B9200S) in PBS, followed by two PBS-T washes and postfixation in 3% formaldehyde (Electron Microscopy Sciences 15714) and 0.1% glutaraldehyde (Electron Microscopy Sciences 16120) for 30 minutes. .. Padlock probe gap-fill and ligation were performed in 1X Ampligase buffer (Lucigen A3210K) containing 50 nM dNTPs (NEB N0447L), 0.1 μM each padlock probe (Integrated DNA Technologies), 200 μg/mL recombinant albumin (NEB B9200S), 0.4 U/μL RNase H (Enzymatics Y9220L), 0.02 U/μL TaqIT polymerase (Enzymatics P7620L), and 0.5 U/μL Ampligase (Lucigen A3210K). .. Samples were incubated in gap-fill and ligation solution at 37 °C for 5 minutes followed by 45 °C for 90 minutes, then washed twice with PBS-T. Rolling circle amplification (RCA) was performed in 1X Phi29 buffer (Thermo Fisher Scientific EP0091) containing 250 μM dNTPs (NEB N0447L), 5% glycerol (MilliporeSigma G5516), 200 μg/mL recombinant albumin (NEB B9200S), and 1 U/μL Phi29 DNA polymerase (Thermo Fisher Scientific EP0091) for 12 hours at 30 °C.
Amplification:Article Title: The one-week automated genome-wide optical pooled screen
Article Snippet: Padlock probe gap-fill and ligation were performed in 1X Ampligase buffer (Lucigen A3210K) containing 50 nM dNTPs (NEB N0447L), 0.1 μM each padlock probe (Integrated DNA Technologies), 200 μg/mL recombinant albumin (NEB B9200S), 0.4 U/μL RNase H (Enzymatics Y9220L), 0.02 U/μL TaqIT polymerase (Enzymatics P7620L), and 0.5 U/μL Ampligase (Lucigen A3210K). .. Samples were incubated in gap-fill and ligation solution at 37 °C for 5 minutes followed by 45 °C for 90 minutes, then washed twice with PBS-T. Rolling circle amplification (RCA) was performed in 1X Phi29 buffer (Thermo Fisher Scientific EP0091) containing 250 μM dNTPs (NEB N0447L), 5% glycerol (MilliporeSigma G5516), 200 μg/mL recombinant albumin (NEB B9200S), and 1 U/μL Phi29 DNA polymerase (Thermo Fisher Scientific EP0091) for 12 hours at 30 °C. .. Samples were washed twice with PBS-T, then 1 μM sequencing primers (Integrated DNA Technologies) were hybridized in 2X SSC (Ambion AM9763) with 10% (v/v) formamide (Thermo Fisher Scientific 4650) for 30 minutes at room temperature and washed twice with PBS-T prior to in situ sequencing.
other:Article Title: Profiling large-scale protein occupancy on bacterial genomes using IPOD-HR.
Article Snippet: Identifying genomic regions bound by individual proteins such as transcription factors is essential to understanding bacterial gene regulation; however, comprehensive understanding of the effect of protein occupancy on gene regulation would be prohibitively laborious and expensive to achieve using methods such as chromatin immunoprecipitation with sequencing (ChIP-seq) and ChIP with exonuclease treatment (ChIP-exo) for every protein and condition of interest.. Here we describe a protocol for performing in vivo protein occupancy display–high resolution (IPOD-HR), a powerful method for genome-wide profiling of protein-bound DNA in prokaryotic systems.. Although assay for transposase-accessible chromatin with sequencing (ATAC-seq) is the method of choice for assaying general protein occupancy in eukaryotic systems, bacterial nucleoid-associated proteins can affect ATAC-seq, rendering it unsuitable for use in bacteria.
Modification:Article Title: The shared genomic history of Middle- to Late-Holocene populations from the Southern Cone of South America.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Ancient skeletal element This study TPGU003, MLP-DA-39, Arg039 Ancient skeletal element This study TPGU004, MLP-DA-46, Arg046 Ancient skeletal element This study TREB001, FCS.TB.E1.1 Ancient skeletal element This study TREB002, FCS.TB.E3.1 Ancient skeletal element This study I20197, MHRTL.ZZ.E1 Ancient skeletal element This study I20341, MHRTL.LM.E1 Ancient skeletal element This study I20342, MHRTL.LTC.E1 Ancient skeletal element This study I20345, MHRTL.LA.E1 Ancient skeletal element This study I20346, MHRTL.B.E1 Chemicals, peptides, and recombinant proteins 0.5 M EDTA, pH 8.0 500 ml Thermo Fisher Scientific AM9261 2-Propanol, 2,5 L, Plastic bottle Merck 1070222511 20% SDS Solution 250 ml Thermo Fisher Scientific AM9820 Adenosine 5’-Triphosphate (ATP) (10 mM) NEB P0756L ATP-Lösung (100 mM) Thermo Fisher Scientific R0441 Bst 2.0 DNA Polymerase NEB M0537L D1000 Sample buffer Agilent 5067-5602 Denhardt′s Solution Merck D9905-5X5ML dNTPs 25 mM Thermo Scientific R1122 DynabeadsTM MyOneTM Streptavidin C1 (10ml) Thermo Fisher Scientific 65002 DynabeadsTM MyOneTM Streptavidin T1 (10 ml) Thermo Fisher Scientific 65602 Ethanol, 2,5 L, Plastic bottle Merck 1009832511 Fast AP (1 U/μl) Thermo Fisher Scientific EF0651 GeneAmpTM 10X PCR Gold Buffer & MgCl2 Thermo Fisher Scientific 4379874 GeneRuler ultra low range DNA ladder Thermo Fisher Scientific SM1211 Guanidine hydrochloride Merck G3272-100G Herculase II 5X rxn buffer Agilent 929677 Herculase II Fusion DNA Polymerase Agilent 600679 Humane Cot-1 DNATM Thermo Fisher Scientific 15279011 Klenow Fragment (10 U/μL) Thermo Fisher Scientific EP0052 Maxima Probe qPCR Mastermix (2x) (1000 rxn) Thermo Fisher Scientific K0262 PB Buffer Qiagen 19066 PEG 8000 Powder, Molecular grade Promega V3011 PfuTurbo Cx Hotstart DNA Polymerase Agilent 600414 Polynucleotide Kinase (PNK) NEB M0201L Proteinase K, Recombinant, PCR-Grade Thermo Fisher Scientific EO0492 Recombinant Albumin Molecular Biology Grade (conc. .. 20 mg/ml) NEB B9200S ROTIPHORESE®50x TAE Buffer, 2.5 l, glass, 1 x 2.5 l Carl Roth CL86.3 Sodium acetate buffer solution, 3 M, pH 5,2 Merck S7899-500ML Sodium chloride solution 5 M Merck S5150-1L Sodium hypochlorite solution, 2.5 l, 12 % Cl, stabilised, technical Carl Roth 9062.4 Sodium phosphat, 0.5 M-Pufferlösung, pH 7.0 Thermo Fisher Scientific J63791.AP SpeedBeads magnetic carboxylate modified particles Merck GE65152105050250 SSC (20X), RNase-frei, 500 ml Thermo Fisher Scientific AM9763 T4 DNA Ligase (5 U/μl) Thermo Fisher Scientific EL0012 T4 DNA Ligase HC (30 U/μl) Thermo Fisher Scientific EL0013 T4 DNA Polymerase 3U/μl NEB M0203L (Continued on next page) ll OPEN ACCESS Current Biology 36, 1–15.e1–e14, May 4, 2026 e2 Article .. REAGENT or RESOURCE SOURCE IDENTIFIER T4 RNA Ligase Reaction Buffer + PEG 8000 + ATP 10 mM NEB B0216L T4-Polynukleotid-Kinase (10 U/μl) Thermo Fisher Scientific EK0032 Tango-Buffer (10X) Life Technologies BY5 TE-Buffer low EDTA, pH 8.0 VWR A8569.0500 TRIS Buffer solution pH 8 (1 mol/l) VWR A4577.1000 TWEEN® 20 Merck P9416-50ML UltraPureTM 1 M Tris-HCI, pH 8,0 Thermo Fisher Scientific 15568025 UltraPureTM Salmon Sperm DNA Solution Thermo Fisher Scientific 15632011 Uracil Glycosylase inhibitor (UGI) NEB M0281L USER Enzyme, recombinant NEB M5505L Water HPLC Plus Merck 34877-2.5L-M EDTA, 05M (pH 8.0) MolecularBiology Grade 100 ml Promega V4231 Proteinase K, Recombinant, PCR-Grade Roche 03115852001 N-Lauroylsarcosine Merck 8147150100 PE Buffer Qiagen 19065 EB Buffer Qiagen 19086 PlatinumTM SuperFi II PCR Master Mix Thermo Fisher Scientific 12368010 Illumina Purification Beads, 100 ml Illumina 20060057 QubitTM 1X dsDNA High Sensitivity (HS) Thermo Fisher Scientific Q33230 Critical commercial assays Amicon® Ultra Zentrifugenfilter, 3 kDa MWCO, 15 ml Merck UFC900308 Amicon® Ultra Zentrifugenfilter, 3 kDa MWCO, 4 ml Merck UFC800308 Biozym Blue S’Green qPCR Kit Separate ROX Biozym 331416XL D1000 ScreenTape Agilent 5067-5582 High Pure Viral Nucleic Acid Large Volume Kit, Roche CVU, Roche 05114403001 MinElute PCR Purification Kit (250) Qiagen 28006 Oligo aCGH/ChIP-on-chip Hybridization Kit Agilent 5188-5220 Quick Ligation Kit NEB M2200L TapeStation Test-Tape (TSTT) Agilent 5067-5601 High Sensitivity D1000 ScreenTape Agilent 5067-5582 High Sensitivity D1000 Reagents Agilent 5067-5583 D1000 ScreenTape Agilent 5067-5584 D1000 Reagents Agilent 5067-5585 15 ml Conical Reservoir Zymo Research C1031-25 Deposited data MtDNA and nuclear DNA sequences (European Nucleotide Archive) This study PRJEB102023 Software and algorithms EAGER version 1.92.38 https://github.com/apeltzer/EAGER-GUI Peltzer et al.124 AdapterRemoval https://github.com/MikkelSchubert/ adapterremoval Schubert et al.125 Burrows-Wheeler-Aligner (BWA) https://github.com/lh3/bwa Li and Durbin126 DeDup https://github.com/apeltzer/DeDup Peltzer et al.124 mapDamage2.0 https://ginolhac.github.io/mapDamage/ Jónsson et al.127 schmutzi https://github.com/grenaud/schmutzi Renaud et al.129 AngsD https://github.com/ANGSD/angsd Korneliussen et al.130 samtools trim https://github.com/samtools/samtools Danecek et al.132 pileupCaller https://github.com/stschiff/sequenceTools N/A (Continued on next page) ll OPEN ACCESS e3 Current Biology 36, 1–15.e1–e14, May 4, 2026 Article
Reverse Transcription:Article Title: The one-week automated genome-wide optical pooled screen
Article Snippet: Samples were washed three times with PBS-T and post-fixed in 3% formaldehyde (Electron Microscopy Sciences 15714) and 0.1% glutaraldehyde (Electron Microscopy Sciences 16120) for 30 minutes, followed by quenching for 1 minute in pH 8 0.2 M Tris-HCl (Thermo Fisher Scientific AM9855G) and three PBS-T washes. .. Reverse transcription was carried out in 1X RevertAid RT buffer (Thermo Fisher Scientific EP0452) containing 250 μM dNTPs (NEB N0447L), 1 μM biotinylated reverse-transcription primer (Integrated DNA Technologies), 200 μg/mL recombinant albumin (NEB B9200S), 0.8 U/μL RiboLock RNase inhibitor (Thermo Fisher Scientific EO0384), and 4.8 U/μL RevertAid H-minus Reverse Transcriptase (Thermo Fisher Scientific EP0452). ..
CtB Assay:Article Title: Dissecting the cellular architecture of breast cancer brain metastases reveals prognostically distinct immune landscapes.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Killing assay: Mouse monoclonal anti-CD28 (clone CD28.2) BioLegend Cat#302902 (RRID:AB_314304) ICB assay: Pembrolizumab Heidelberg University Hospital Pharmacy NA ICB assay: Tiragolumab Hycultec Cat# HY-P9986 (RRID:AB_3695514) Biological samples BCBM tissue specimens Biobank of the Department of Neurosurgery, Heidelberg University Hospital, Heidelberg, Germany N/A Blood samples of BCBM patients Biobank of the Department of Neurosurgery, Heidelberg University Hospital, Heidelberg, Germany N/A Chemicals, peptides, and recombinant proteins 2-Methylbutane Carl Roth Cat#3927.2 AMPure XP beads Beckman Coulter Cat#A63881 Biocoll solution Biochrom Cat#L6115 Bovine serum albumin (BSA), molecular biology grade New England Biolabs Cat#B9000S Brefeldin A Solution Biolgend Cat#420601 Buffer EB Qiagen Cat#19086 Calcium chloride (CaCl2) 1 M Hölzel Diagnostika Cat#103047-1 CellTiter-Blue assay Promega Cat#G8081 DAKO diluent, background reducing Agilent Dako Cat#S302283-2 DAPI (4’,6-Diamidin-2-phenylindol) Thermo Fisher Scientific Cat#D1306 Deoxyribonuclease I (DNAse I) Sigma-Aldrich Cat#DN25-100MG EGF Novoprotein Cat#C029 β-Estradiol Sigma-Aldrich Cat#E2758 bFGF Novoprotein Cat#C046 BIT admixture 100 PELOBiotech Cat#PB-SH-033-0000 DMEM/F-12, no phenol red Thermo Fisher Scientific Cat#21041025 DMEM, high glucose, GlutaMAX supplement Thermo Fisher Scientific Cat#61965059 Dulbecco’s Phosphate Buffered Saline (DPBS) Thermo Fisher Scientific Cat#14190-094 EDTA (Versen) 1% in PBS Genaxxon Bioscience Cat#C4263.0100 FCS (Fetal calf serum) Sigma-Aldrich Cat#S0615 Fixable Viability Dye eFluor 506 Thermo Fisher Scientific Cat#65-0866-14 Fixable Viability Dye eFluor 450 Thermo Fisher Scientific Cat#65-0863-14 GlutaMAX Supplement Thermo Fisher Scientific Cat#35050038 Glycerol Sigma-Aldrich Cat#1040950250 IgG from human serum Sigma-Aldrich Cat#I2511 IL-2, human, recombinant Stemcell Technologies Cat#78036.3 IncuCyte Caspase-3/7 Green Dye Sartorius Cat#4440 IncuCyte Cytolight Rapid Red Dye Sartorius Cat#4706 ImmunoCult Human CD3/CD28/CD2 T Cell Activator Stemcell Technologies Cat#10970 ImmunoCult-XF T Cell Expansion Medium Stemcell Technologies Cat#10981 Liberase DH Research Grade Merck Cat#05401089001 Methylene blue Conatex Cat#9993339 Magnesium chloride (MgCl2) 1 M Sigma Aldrich Cat#M1028 Mowiol 4-88 Carl Roth Cat#0713.2 (Continued on next page) ll OPEN ACCESSArticle Cancer Cell 44, 1–22.e1–e16, June 8, 2026 e3 .. REAGENT or RESOURCE SOURCE IDENTIFIER Nuclease-free water Thermo Fisher Scientific Cat#AM9937 Penicillin-Streptomycin (10.000 U/ml) Thermo Fisher Scientific Cat# 15140122 Propidium Iodide BioLegend Cat#421301 Sodium chloride (NaCl) 5 M Thermo Fisher Scientific Cat#AM9760G Staurosporine Hycultec Cat# HY-15141 Synth-a-Freeze cryopreservation medium Thermo Fisher Scientific Cat#A1254201 Tissue-Tek O.C.T. compound Science Services Cat#SA6255012 Tris (pH 7.5), 1 M Thermo Fisher Scientific Cat#15567027 Tween-20 Sigma-Aldrich Cat#P-7949 Critical commercial assays Chromium NEXT GEM Chip K Single Cell Kit 10x Genomics Cat#1000286 and Cat#1000287 Chromium Next GEM Single Cell 5’ Kit v2 - Chromium Next GEM Single Cell 5’ Gel Bead Kit v2 - Chromium Next GEM Single Cell 5ʹ GEM Kit v2 - Library Construction Kit 10x Genomics Cat#1000263 - Cat#1000264 and Cat#1000267 - Cat#1000244 and Cat#1000266 - Cat#1000190 and Cat#1000196 Tube, Dynabeads MyOne SILANE 10x Genomics Cat#2000048 Dual Index Kit TT Set A 10x Genomics Cat#1000215 EasySep Human CD3 Positive Selection Kit II Stemcell Technologies Cat#17851 Foxp3 / Transcription Factor Staining Buffer Set Thermo Fisher Scientific Cat# 00-5523-00 IC fixation buffer Thermo Fisher Scientific Cat# 00-8222-49 Infinium OncoArray-500K BeadChip Illumina Cat#WG-355-1001 LIVE/DEAD Cell Imaging Kit (488/570) Thermo Fisher Scientific Cat#R37601 Tumor Dissociation Kit, human Miltenyi Biotec Cat#130-095-929 Deposited data Bulk RNA-seq data, BCBM discovery cohort This paper German Human Genome-Phenome Archive (GHGA) as GHGAD28055171876192 Gene Expression Omnibus: GSE322943 Bulk RNA-seq data, BCBM CATCH cohort This paper German Human Genome-Phenome Archive (GHGA) as GHGAD51052817169855 Single nuclei RNA-seq data, BCBM This paper Zenodo at 10.5281/zenodo.17723784 German Human Genome-Phenome Archive (GHGA) as GHGAD48917471237543 Gene Expression Omnibus: GSE324453 Spatial transcriptomics data This paper Zenodo at https://doi.org/10.5281/zenodo.
Saline:Article Title: Dissecting the cellular architecture of breast cancer brain metastases reveals prognostically distinct immune landscapes.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Killing assay: Mouse monoclonal anti-CD28 (clone CD28.2) BioLegend Cat#302902 (RRID:AB_314304) ICB assay: Pembrolizumab Heidelberg University Hospital Pharmacy NA ICB assay: Tiragolumab Hycultec Cat# HY-P9986 (RRID:AB_3695514) Biological samples BCBM tissue specimens Biobank of the Department of Neurosurgery, Heidelberg University Hospital, Heidelberg, Germany N/A Blood samples of BCBM patients Biobank of the Department of Neurosurgery, Heidelberg University Hospital, Heidelberg, Germany N/A Chemicals, peptides, and recombinant proteins 2-Methylbutane Carl Roth Cat#3927.2 AMPure XP beads Beckman Coulter Cat#A63881 Biocoll solution Biochrom Cat#L6115 Bovine serum albumin (BSA), molecular biology grade New England Biolabs Cat#B9000S Brefeldin A Solution Biolgend Cat#420601 Buffer EB Qiagen Cat#19086 Calcium chloride (CaCl2) 1 M Hölzel Diagnostika Cat#103047-1 CellTiter-Blue assay Promega Cat#G8081 DAKO diluent, background reducing Agilent Dako Cat#S302283-2 DAPI (4’,6-Diamidin-2-phenylindol) Thermo Fisher Scientific Cat#D1306 Deoxyribonuclease I (DNAse I) Sigma-Aldrich Cat#DN25-100MG EGF Novoprotein Cat#C029 β-Estradiol Sigma-Aldrich Cat#E2758 bFGF Novoprotein Cat#C046 BIT admixture 100 PELOBiotech Cat#PB-SH-033-0000 DMEM/F-12, no phenol red Thermo Fisher Scientific Cat#21041025 DMEM, high glucose, GlutaMAX supplement Thermo Fisher Scientific Cat#61965059 Dulbecco’s Phosphate Buffered Saline (DPBS) Thermo Fisher Scientific Cat#14190-094 EDTA (Versen) 1% in PBS Genaxxon Bioscience Cat#C4263.0100 FCS (Fetal calf serum) Sigma-Aldrich Cat#S0615 Fixable Viability Dye eFluor 506 Thermo Fisher Scientific Cat#65-0866-14 Fixable Viability Dye eFluor 450 Thermo Fisher Scientific Cat#65-0863-14 GlutaMAX Supplement Thermo Fisher Scientific Cat#35050038 Glycerol Sigma-Aldrich Cat#1040950250 IgG from human serum Sigma-Aldrich Cat#I2511 IL-2, human, recombinant Stemcell Technologies Cat#78036.3 IncuCyte Caspase-3/7 Green Dye Sartorius Cat#4440 IncuCyte Cytolight Rapid Red Dye Sartorius Cat#4706 ImmunoCult Human CD3/CD28/CD2 T Cell Activator Stemcell Technologies Cat#10970 ImmunoCult-XF T Cell Expansion Medium Stemcell Technologies Cat#10981 Liberase DH Research Grade Merck Cat#05401089001 Methylene blue Conatex Cat#9993339 Magnesium chloride (MgCl2) 1 M Sigma Aldrich Cat#M1028 Mowiol 4-88 Carl Roth Cat#0713.2 (Continued on next page) ll OPEN ACCESSArticle Cancer Cell 44, 1–22.e1–e16, June 8, 2026 e3 .. REAGENT or RESOURCE SOURCE IDENTIFIER Nuclease-free water Thermo Fisher Scientific Cat#AM9937 Penicillin-Streptomycin (10.000 U/ml) Thermo Fisher Scientific Cat# 15140122 Propidium Iodide BioLegend Cat#421301 Sodium chloride (NaCl) 5 M Thermo Fisher Scientific Cat#AM9760G Staurosporine Hycultec Cat# HY-15141 Synth-a-Freeze cryopreservation medium Thermo Fisher Scientific Cat#A1254201 Tissue-Tek O.C.T. compound Science Services Cat#SA6255012 Tris (pH 7.5), 1 M Thermo Fisher Scientific Cat#15567027 Tween-20 Sigma-Aldrich Cat#P-7949 Critical commercial assays Chromium NEXT GEM Chip K Single Cell Kit 10x Genomics Cat#1000286 and Cat#1000287 Chromium Next GEM Single Cell 5’ Kit v2 - Chromium Next GEM Single Cell 5’ Gel Bead Kit v2 - Chromium Next GEM Single Cell 5ʹ GEM Kit v2 - Library Construction Kit 10x Genomics Cat#1000263 - Cat#1000264 and Cat#1000267 - Cat#1000244 and Cat#1000266 - Cat#1000190 and Cat#1000196 Tube, Dynabeads MyOne SILANE 10x Genomics Cat#2000048 Dual Index Kit TT Set A 10x Genomics Cat#1000215 EasySep Human CD3 Positive Selection Kit II Stemcell Technologies Cat#17851 Foxp3 / Transcription Factor Staining Buffer Set Thermo Fisher Scientific Cat# 00-5523-00 IC fixation buffer Thermo Fisher Scientific Cat# 00-8222-49 Infinium OncoArray-500K BeadChip Illumina Cat#WG-355-1001 LIVE/DEAD Cell Imaging Kit (488/570) Thermo Fisher Scientific Cat#R37601 Tumor Dissociation Kit, human Miltenyi Biotec Cat#130-095-929 Deposited data Bulk RNA-seq data, BCBM discovery cohort This paper German Human Genome-Phenome Archive (GHGA) as GHGAD28055171876192 Gene Expression Omnibus: GSE322943 Bulk RNA-seq data, BCBM CATCH cohort This paper German Human Genome-Phenome Archive (GHGA) as GHGAD51052817169855 Single nuclei RNA-seq data, BCBM This paper Zenodo at 10.5281/zenodo.17723784 German Human Genome-Phenome Archive (GHGA) as GHGAD48917471237543 Gene Expression Omnibus: GSE324453 Spatial transcriptomics data This paper Zenodo at https://doi.org/10.5281/zenodo.
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