METADATA
last updated: 2026-02-15 RT initial conversion
file_name: Assay and Reagent Manufacturing Diagrams.md
file_date: 2022-02-01
title: FloodLAMP Assay and Reagent Manufacturing Diagrams
category: guides
subcategory: manufacturing
tags: 
source_file_type: gslide
xfile_type: pptx
gfile_url: https://docs.google.com/presentation/d/10kiok18TcbatUeareNyUyrcr90_ib0gT56NpWsIh58U
xfile_github_download_url: https://raw.githubusercontent.com/FocusOnFoundationsNonprofit/floodlamp-archive/main/guides/manufacturing/Assay%20and%20Reagent%20Manufacturing%20Diagrams.pptx
pdf_gdrive_url: https://drive.google.com/file/d/1uyKevLkDpU1De9Gl8Y2fi1CHgNfvMl7f
pdf_github_url: https://github.com/FocusOnFoundationsNonprofit/floodlamp-archive/blob/main/guides/manufacturing/Assay%20and%20Reagent%20Manufacturing%20Diagrams.pdf
conversion_input_file_type: pptx
conversion: msmid
license: CC BY 4.0 - https://creativecommons.org/licenses/by/4.0/
tokens: 1958
words: 925
notes: This is a presentation (pptx/Google Slides) consisting primarily of manufacturing process flow diagrams (images). The markdown content lists slide titles only as placeholders since the slides are image-based diagrams. Refer to the source PDF or pptx for the actual diagrams.
summary_short: The FloodLAMP Assay and Reagent Manufacturing Diagrams is a 19-slide presentation containing process flow diagrams for all key reagent manufacturing and assay workflows. It covers the FloodLAMP Direct RNA Assays (QuickColor LAMP and EasyPCR) at 48- and 96-reaction scales, validation first-run setup, gamma contrived positive preparation, 100X Inactivation Solution manufacturing (standard and XL batches), 1X Inactivation Saline Solution preparation, Twist Positive Control aliquoting, Gamma Blue and 10K dilution series, Primer-Guanidine Solution (PGS) batching, colorimetric CLAMP assay setup, 5X PCR primer preparation, and PCR assay reaction mix preparation.


CONTENT

## Slide 1: FloodLAMP Direct RNA Assays - 48 Reaction Volumes
_Manufacturing process flow diagram showing reagent preparation and assay workflow for 48-reaction scale. Covers 100X Inactivation Solution, 0.9% Saline, Primer-Guanidine Solution (PGS, 550µL), Colorimetric LAMP MM (655µL), PCR Reaction Mix, and both QuickColor LAMP Test (65°C/25min) and EasyPCR Test (qPCR, 80min) pathways._

## Slide 2: FloodLAMP Direct RNA Assays - 96 Reaction Volumes
_Manufacturing process flow diagram showing reagent preparation and assay workflow for 96-reaction scale. Same reagent pathways as Slide 1 but with scaled volumes: PGS (1080µL), Colorimetric LAMP MM (1286µL), CLAMP Reaction Mix (2366µL, 25µL/rxn)._

## Slide 3: Validation: First Run - 24 Reaction Volumes
_Validation run diagram for 24-reaction scale with both CLAMP (PGS 275µL, CLAMP MM 327µL, 602µL total RM) and PCR (5X PCR Primers 105µL, PCR MM 263µL, RT 26.3µL) assay pathways. Includes positive and negative controls in the 24-tube sample plate._

## Slide 4: Gamma Contrived Positive Diagram (v1.3)
_Process flow for preparing Gamma Contrived Positive (GCP) samples. Shows dilution from Gamma Blue Dilution (GBD, 1000 cp/µL) using anterior nares (AN) swabs dried >20 min, with 1X Inactivation Saline Solution elution and standard inactivation procedures._

## Slide 5: 100X Inactivation Solution XL Diagram
_Extended-scale manufacturing diagram for 100X Inactivation Solution. Key source materials: TCEP (25g), 10N NaOH (1L), 0.5M EDTA (100mL). Includes component volume calculation table with Standard and Adjusted columns, plus 0.5M TCEP intermediate preparation step._

## Slide 6: 1X Inactivation Saline Solution Diagram
_Process flow for preparing 1X Inactivation Saline Solution (1XISS). Combines 5M NaCl-derived 0.9% Saline with 100X Inactivation Solution at ~1/100th volume. Shows 50mL Falcon tube aliquoting and reference to Saline Batch Record v1.0 and Inactivation Protocol v5.1._

## Slide 7: Twist Positive Control Diagram
_Aliquoting workflow for Twist Positive Controls (TPC). Key source: Twist 102019 (~1e6 cp/µL, 100µL) and Total Human RNA (50ng/µL, 100µL). Creates TSPC (5µL x 19), TRPC source stock (1000 cp/µL, 490µL x 10), TFPC (20µL x 24), and TPC working aliquots (125 cp/µL, 10µL x 16). All stored at -80°C._

## Slide 8: Gamma Blue Dilution (GBD) Diagram
_Dilution workflow from BEI gamma-irradiated SARS-CoV-2 (NR-52287, 1.75e6 cp/µL). Creates GBEI aliquots (5µL x 100 in PCR strip tubes), then dilutes to GBD (1000 cp/µL) in 30mL Chub (8.75mL), and final aliquots in 1.5mL tubes (83 x 105µL). Reference: Gamma Blue Dil Batch Record v1.1._

## Slide 9: Gamma 10K Dilution (G10K) Diagram
_Dilution workflow from BEI source (1.75e6 cp/µL) to 10,000 cp/µL working stock. Creates GBEI aliquots (5µL x 100), then G10K in 1.5mL screw cap (875µL at 10,000 cp/µL), and final aliquots (32 x 27µL). Reference: Gamma 10K Dil Batch Record vX._

## Slide 10: Primer-Guanidine Solution (PGS) Diagram
_Manufacturing diagram for PGS batches. Combines dry LAMP primer sets (As1e, N2, E1) resuspended in NFWater with 6M Guanidine-HCl (200µL) to create 10X LAMP Primer Mix, then dilutes to PGS. Aliquot options: 92µL x 136 (8 rxn), 275µL x 45 (24 rxn), or 1120µL x 11 (96 rxn). PGS specs: Primers 2.38X, GUD 95.1nM. Reference: Batch Record v2.0._

## Slide 11: Colorimetric Assay Diagram - 24 and 96 Reactions (CLAMP Assay v2.5)
_CLAMP reaction mix preparation for 24-rxn (PGS 275µL + CLAMP MM 327µL = 602µL RM, 73µL/tube fill strips) and 96-rxn (PGS 1080µL + CLAMP MM 1286µL = 2366µL RM, 290µL/tube fill strips) scales. Each CLAMP reaction: 23µL MM + 2µL sample._

## Slide 12: Colorimetric Assay Diagram - 48 Reactions (CLAMP Assay v2.5)
_CLAMP reaction mix preparation for 48-reaction scale. Uses PGS and Colorimetric LAMP MM from frozen storage. Fill strip volumes at 146µL/tube. Note: some volume fields marked "FIX" in original._

## Slide 13: 5X PCR Primers Diagram
_Manufacturing diagram for 5X PCR Primer working stocks. Combines source primers with 11.562mL NFWater to create 12.5mL bulk in 30mL Chub. Aliquots to 1.5mL tubes (105.6µL x 10, 24 rxn, 10% over) and 5mL tubes (1.1mL x 11). References: 5X PCR Primer Source Stock v1.1 and Working Stock v1.0._

## Slide 14: PCR Assay Diagram (PCR Assay v1.3)
_PCR reaction mix preparation for 24-reaction scale. Combines Luna PCR MM (from NEB E3006 Kit), LUNA RT (26.4µL from 100µL aliquot), 5X PCR Primers (105.6µL), and NFWater (79.2µL) into 472.5µL total RM in 1.5mL tube. Fill strip at 57µL/well. Each PCR reaction: 18µL MM + 2µL sample._

## Slide 15: EXTRA
_Blank/placeholder slide._

## Slide 16: 100X Inactivation Solution Diagram (Standard)
_Standard-scale manufacturing diagram for 100X Inactivation Solution. Shows preparation of intermediate stocks: 0.5M TCEP (in 250mL glass bottle + 100mL NFWater), 3.83N NaOH (in 250mL glass bottle + 74mL NFWater), and 0.5M EDTA. Aliquoting to 30mL Chubs and 15mL Falcons, then combining into 100XIS (TCEP 0.25M, EDTA 0.1M, NaOH 1.15N) in 30mL Chub, with final 1mL x 24 aliquots in 1.5mL tubes at -20°C. References batch records for TCEP v1.1, NaOH v1.0, EDTA v1.0, and 100XIS v1.0._

## Slide 17: FloodLAMP Overview - Streamlined Sample Prep
_Overview diagram showing the complete FloodLAMP workflow: 100X Inactivation Solution → 1X Inactivation Saline Solution → sample processing → dual test paths. QuickColor LAMP Test (high sensitivity 90%, ultra-high throughput, ideal for serial screening). EasyPCR Test (very high sensitivity 98%, medium throughput 1.5 hours/94, ideal for diagnostics/reflex/confirm). Highlights streamlined sample prep with upfront swab pooling and same sample for both tests._

## Slide 18: Streamlined Sample Prep (Simplified)
_Simplified version of the overview diagram showing the three main workflow blocks: Streamlined Sample Prep → QuickColor LAMP Test → EasyPCR Test._

## Slide 19: FloodLAMP Lab Playbook
_Diagram showing reflex testing workflow: Positives or Invalids from QuickColor COVID-19 Test reflexed using FloodLAMP EasyPCR COVID-19 Test._